Stephen N. Rudnick

James J. McDevitt

Melvin W. First

John D. Spengler

Air Transportation Center of Excellence for 

Airliner Cabin Environment Research

Harvard School of Public Health

Boston, MA 02115

April 2009

Final Report

Inactivating Influenza Viruses on 

Surfaces Using Hydrogen Peroxide 

or Triethylene Glycol at Low Vapor 

Concentrations

DOT/FAA/AM-09/7

Office of Aerospace Medicine

Washington, DC 20591

OK-09-0434-JAH

Federal Aviation

Administration
NOTICE

This document is disseminated under the sponsorship 

of the U.S. Department of Transportation in the interest 

of information exchange. The United States Government 

assumes no liability for the contents thereof.

___________

This publication and all Office of Aerospace Medicine 

technical reports are available in full-text from the Civil 

Aerospace Medical Institute’s publications Web site: 

www.faa.gov/library/reports/medical/oamtechreports


Technical Report Documentation Page 

1. Report No. 2. Government Accession No. 3. Recipient's Catalog No. 

DOT/FAA/AM-09/7 

4. Title and Subtitle 5. Report Date 

Inactivating Influenza Viruses on Surfaces Using Hydrogen Peroxide or April 2009 

Triethylene Glycol at Low Vapor Concentrations 6. Performing Organization Code 

 7. Author(s) 8. Performing Organization Report No. 

Rudnick SN, McDevitt JJ, First MW, Spengler JD 9. Performing Organization Name and Address 10. Work Unit No. (TRAIS) 

 11. Contract or Grant No. 

Air Transportation Center of Excellence for Airliner 

Cabin Environment Research 

Harvard School of Public Health 

665 Huntington Avenue 

Boston, MA 02115 

 

12. Sponsoring Agency name and Address 13. Type of Report and Period Covered 

 

 

 

Office of Aerospace Medicine 

Federal Aviation Administration 

800 Independence Ave., S.W. 

Washington, DC 20591 14. Sponsoring Agency Code 15. Supplemental Notes 

Work was accomplished under Public law 108-76 16. Abstract Any of the exposed surfaces in airplanes can become contaminated with infectious viruses, such as influenza, 

and facilitate transmission of disease. In this study, we disinfected surfaces contaminated with influenza 

viruses. Selection of the specific decontamination agents used in this study was based on three criteria: 1) no 

harm would be caused to the mechanical components or avionics of the airplane, 2) no potentially harmful 

residue would be left behind, and 3) the airplane could quickly be returned to service. We chose two 

decontamination agents that we believed fulfilled these criteria: 1) hydrogen peroxide (HP) at vapor 

concentrations in air below 100 ppm and 2) triethylene glycol (TEG) saturated air, which has a TEG vapor 

concentration of about 2 ppm at 25°C. For influenza viruses deposited on stainless-steel coupons and 

exposed for 15 min to 10 to 90 ppm of HP vapor, the number of log reductions of active viruses ranged 

from 3.6 to 4.7. The number of log reductions, however, was not linear with time; log reduction rate 

decreased significantly with increasing exposure time. For example, at a HP vapor concentration of 57 ppm, 

the number of log reductions was 3.2 after 2.5 min but just 4.0 after 10 min. Even after 60 min, the number 

of log reductions was only 5.6. At a HP vapor concentration of 10 ppm, the number of log reductions was 

2.0 after 2.5 min. This corresponds to 99% inactivation of viruses, a significant reduction for such a low HP 

vapor concentration. For air saturated with TEG at 25-29°C, the number of log reductions of influenza 

viruses versus exposure time followed a linear relationship reasonably well. The decontamination rate was 

equal to 1.3 log reductions per hour. The potential for damage to the mechanical components or avionics of 

the airplane at a TEG vapor concentration of 2 ppm would be expected to be minimal. In addition, at a 2 

ppm TEG vapor concentration, there is essentially no health risks to people. 

 

17. Key Words 18. Distribution Statement 

 

 

Surface Decontamination, Hydrogen Peroxide Vapor, 

Vaporized Hydrogen Peroxide, Triethylene Glycol Vapor, 

Influenza Virus

Document is available to the public through the 

Defense Technical Information Center, Ft. 

Belvoir, VA 22060; and the National Technical 

Information Service, Springfield, VA 22161 19. Security Classif. (of this report) 20. Security Classif. (of this page) 21. No. of Pages 22. Price 

Unclassified Unclassified 14 

Form DOT F 1700.7 (8-72) Reproduction of completed page authorized 

ACKNOWLEDGMENTS

The authors thank Dr. William Gale for providing guidance during this project and for his assi

procuring stainless-steel surfaces used during experiments. We also thank the technicians who work

this project and contributed to its completion: Gihan Amarasiriwardena, Kenneth Chin, Alissa 

Christa James, and Chika Uchiyam

ABBREVIATIONS

As used in this report, the following abbreviations/acronyms have the meanings indicated

Abbreviation Meani

ACGIH  - - - American Conference of Governmental Industrial H

BSA- - - - Bovine serum 

DPBS - -- -- -- Dulbecco’s phosphate buffered salin

DPBS++- - - - DPBS with calcium and m

EPA  -- -- -- -- Environmental Pr

FFU  -- -- -- -- Fluoresce

HP  - - - Hydrogen pe

MDCK  - - - Madin-Darby canin

OSHA  -- -- -- Occupational Safety and Health 

PEL  -- -- -- -- Permissib

RH  - - - Relative 

SARS  - - - Severe acute respiratory sy

TEG - - Triethylene gly

TLV®- - - - Threshold Limi

UV  - - - Ultra

UVGI  -- -- -- Ultraviolet germic

MATHEMATICAL SYMBOLS

As used in this report, the following symbols have the meanings indicated

Symbol Meani

f - - Fraction of viruses remaining

logU- - - - Mean of the logarithm

Uo log- -- -- -- Mean of the logar

n- - - - Number of log redu

n- - - - Average number of log redu

pTEG- -- -- -- -- Partial pressu

P- - - - Ambient pressure o PTEG - - Vapor pressure of pure l

U slog- -- -- -- -- Standard dev

Uo slog - - Standard deviation of lo

n s- -- -- -- -- -- Standard deviation

T  -- -- -- -

U- -- -- -- -- -- Number of FFU per volume of rinsate from 

Uo- -- -- -- -- -- -- -- -- -- Number of FFU per volume of rinsate from an unexp

TEG y - - TEG mole fraction in the gas p

v

C

INTRODUCTION  

Background. 

Hydrogen Peroxide Vapor Decontaminatio

Triethylene Glycol Vapor Decontaminatio

METHOD AND MATERIALS. 

Influenza Virus.

Preparation and Treatment of Test Surfaces. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 

Exposure Chamber and Test Methodology. 

Calculations.

RESULTS. 

Hydrogen Peroxide Vapor Decontaminatio

Triethylene Glycol Vapor Decontaminatio

Viability Tests at Ambient Condit

DISCUSSION. 

Hydrogen Peroxide Vapor Decontaminatio

Triethylene Glycol Vapor Decontaminatio

CONCLUSIONS. 

REFERENCES. 

Inactivating Influenza Viruses on Surfaces Using Hydro

or Triethylene Glycol at Low Vapor Con

INTRODUCTION 

Background

Preventing the spread of smallpox, influenza, SARS

and other virus-caused diseases on commercial airpla

and other public venues is a significant challenge 

the public health community (Musher 2003; Olsen

Chang, Cheung, et al. 2003). Transfer of viruses from

an infected individual to an uninfected one can o

through various modes of transmission: 1) by dir

contact, 2) via fomites (inanimate objects capable

carrying infectious viruses), 3) from the direct s

of large droplets from an infected person, and 4) from

droplet nuclei, which are very small dried droplets th

can stay suspended in the air for long periods of t

(Roy & Milton 2004). In this study, we evaluated th

efficacy of various relatively gentle methods for decontaminating fom

Any of the exposed surfaces in airplanes or othe

vehicles used for public transportation can become contaminated with infectious viruses and be respons

disease transmission. In this study, we disinfected sur

contaminated with influenza A viruses, whose subty

may have the potential to cause a pandemic propagate

worldwide by commercial travel. It may not be necessar

to sterilize an airplane cabin; significant reduction

potential for disease transmission would be benefic

Selection of specific decontamination methods u

in this study was based primarily on three criteria: 1

method would not be expected to cause damage to the 

mechanical components or avionics of the airplane, 2) 

method would leave no potentially harmful residue, and 3

the method would require a relatively brief period of t

so that an airplane or other means of public transportati

could be put back into service quickly. We chose thr

methods that fulfill these criteria: 1) relatively low vap

concentrations (<100 ppm) of hydrogen peroxide (HP)

2) very low vapor concentrations of triethylene glyco

(TEG), and 3) thermal decontamination using heat

air. The efficacy of the decontaminants HP and TE

is the subject of this report. A separate report discuss

thermal decontamination. As a baseline for compari

and because of its importance, the length of time th

influenza viruses remain active on surfaces under amb

conditions was also determine

We eliminated many other decontamination meth

because they did not adhere to the desired criteria. Specifically, methods that rely on chlorine dioxide, formaldehyde, ethylene oxide, and methyl bromide we

likely unacceptable due to their potential for damage 

the airplane and their toxicity to humans. In addi

despite its effectiveness for inactivating viruses and bacteria, we did not choose ultraviolet germicid

(UVGI) because viruses can be protected from UV rays

by lurking in the shadows and because the UV rays ar

not very penetrating, allowing a coating of dust or oth

material to protect viruses from the UV rays. UVGI 

more appropriate for air decontamination (First, Rud

Banahan, et al. 2007; Rudnick & First 2007

Hydrogen Peroxide Vapor Decontamination

We were unable to find any peer-reviewed publications on surface decontamination of influenza v

using HP at vapor concentrations below 100 ppm. I

the few published studies on surface decontamination

influenza viruses at higher HP concentrations, the d

virus suspension was exposed to a relatively large do

of HP vapor; that is, the HP vapor concentration wa

relatively high and exposure time relatively long. N

of these studies gave results on virus inactivation ve

dose. For example, the effect of HP vapor on surfacedeposited influenza viruses was evaluated by Hecker

Best, Jordan, et al. (1997) at a HP vapor concentration

of about 1200 ppm and exposure time of about 30 min

Although Heckert, Best, Jordan, et al. showed an overall 

reduction of influenza viruses of about 6 logs, wh

was their limit of detection, only about 3 logs were attributable to HP vapor; the remainder was due to

of drying at ambient conditions and heat exposure

30-40°C. Other studies using influenza as the challen

viruses had similar limitations. In a recent review a

De Benedictis, Beato, and Capua (2007) conclude tha

“reports on the specific efficacy against avian influe

viruses of hydrogen peroxide are contradictory, and f

this reason additional information on its viricidal

is necessary.

Triethylene Glycol Vapor Decontamination

Although we were unable to find any publications o

the use of TEG vapor to decontaminate surfaces, it ha
been used to disinfect air. Although other glycols can al

be used to disinfect air (Robertson, Bigg, Miller, et

1941;Robertson, Loosli, Puck, et al. 1941), TEG vapor

is the most suitable because of its extremely low vap

pressure, which results in very low air concentratio

In addition, TEG in aerosol form is commonly used 

theatrical “smoke” such as in venues for Broadway productions (Burr, Van Gilder, Trout, et al. 1994). As a result

this and other properties, TEG vapor is believed to do

harm to humans (EPA 2005a) or damage to environmental

surfaces (Lester, Kaye, Robertson, et al. 1950). There are 

a large number of journal publications, primarily f

the 1940s, on the use of TEG vapor for air disinfecti

TEG vapor has been shown to exert lethal action agains

a wide variety of airborne infectious agents inc

bacteria, viruses, and fungi (Lester, Kaye, Robertso

et al. 1950). In particular, TEG vapor was found to be

an effective decontaminant agent for airborne influe

viruses (Robertson, Puck, Lemon, et al. 1943).

METHOD AND MATERIALS

Influenza Virus

Virus Stock. A frozen suspension of influenza virus

(A/PR/8/34 H1N1) was purchased from Advanced Biotechnologies (Columbia, MD). It was thawed, div

into single-use packets, refrozen, and stored at −80°

until needed.

Virus Assay. A fluorescent focus reduction assay

(Hartshorn, White, Tecle, et al. 2007) was used to

measure the titer of virus suspensions before and aft

decontamination. Confluent monolayers of Madin-Dar

canine kidney (MDCK) cells were prepared in 96-we

plates. Each well was inoculated with 50 μL of coupo

rinsate (or serial dilutions of the rinsate) and incu

at 37°C for 45 min in a 5% C

 environment. After

washing the infected cells using assay media compo

of Dulbecco’s Modified Eagle’s Medium (Mediatec

Herndon, VA) with 0.1% bovine serum albumin (BS

(SeraCare, Milford, MA), the cells were incubated for 

h at 37°C in a CO

 environment. After incubation, t

cells were washed three times with Dulbecco’s phosphat

buffer saline with calcium and magnesium (DPBS

(Hyclone Laboratories, Logan, UT) and fixed with a

aqueous solution of 80% acetone for 10 min at 4°C

The infected MDCK cells were then labeled for 30 mi

at 4°C with 50 μL of nucleoprotein antibody soluti

which was made by adding 50 μL of mouse monoclona

antibodies (Centers for Disease Control, Atlanta, G

catalog #VS2366) to 5 mL of Dulbecco’s phosphate 

buffer saline (DPBS) (Hyclone Laboratories, Logan

UT) containing 1% BSA, 1% heat inactivated hu

serum (Mediatech, Herndon, VA), and 0.02% sodiu

azide. After washing three times with DPBS, the ce

were incubated with tagging solution, which was 

by adding 50 μL of rhodamine-labeled goat anti-mou

IgG (Jackson ImmunoResearch Laboratories, West Grove,

PA, catalog #115026062) to 5 mL of DPBS containin

1% BSA, 1% heat inactivated human serum, and 0.02

sodium azide. The number of cells having a fluoresce

foci, which are referred to as fluorescent focus units (FFU

were then counted using an Olympus CKX-41 inverte

fluorescent microscope (Olympus, Center Valley, PA).

Based on this assay, the titer of the single-use packets 

influenza virus suspension after being thawed was ab

109 FFU/mL.

Preparation and Treatment of Test Surfaces

One-inch by three-inch stainless-steel coupons we

used as test surfaces. For tests using higher concentrations of HP vapor or TEG vapor, 50 μL of influenza v

suspension was seeded onto a predetermined number o

coupons. For lower decontaminant concentrations, th

influenza virus suspension was diluted prior to 

seeded onto coupons. All of the coupons were placed 

into a biological safety cabinet until the deposited

had evaporated. The required drying time was betwe

20 and 30 min, depending on ambient conditi

Some of these seeded coupons, which are referred to

as “control” coupons, were left in the biological safe

cabinet, where they continued to be exposed to filtere

room air at ambient conditions. The remaining s

coupons (along with a clean coupon used as a negativ

control) were placed in an exposure chamber containi

HP vapor or TEG vapor. 

The coupons were removed from the exposure chamber 

after predetermined exposure times. The control coupons that were left in the biological safety cabine

no exposure to HP or TEG and, thus, were considered

to be unexposed coupons. Immediately after the last

coupon was removed from the exposure chamber, each 

of the seeded coupons was rinsed with DPBS++ usi

the following procedure: The clearly marked portion o

the coupon where the viruses had been initially de

ited was rinsed 25 times with a single 500-μL porti

DPBS++ using a pipette. No visible residue remain

fluorescent focus reduction assay was then done on the

rinsate and/or diluted rinsate from each coupon. On

about 50% of the influenza viruses that were seeded ont

the control slides were recovered

Exposure Chamber and Test Methodology

Hydrogen Peroxide Vapor Tests. Tests in whic

influenza viruses deposited on stainless-steel coup

were exposed to HP vapor were done in a 130-L cubical plexiglass chamber located within a laboratory 
hood. A shallow pool of an aqueous solution of 35%

HP (VWR, West Chester, PA), diluted with water t

a predetermined HP concentration that was calculate

to provide the desired HP vapor concentration, cover

much of the floor area of the chamber. Prediction of th

HP concentration in an aqueous solution required fo

specific HP vapor concentration was based on publish

correlations (Schumb, Satterfield & Wentworth 1955).

The air inside the chamber was kept well mixed thro

the use of two small fans. To maintain the desired relative humidity (RH), 17 L/min of dry air was a

the chamber. Temperature and RH were monitored and

recorded every 30 s using a HOBO (Onset Computer

Corp., Buzzards Bay, MA). RH and temperature were also 

measured periodically with a hygrometer (Omega Engineering, Stamford, CT) and mercury thermom

HP vapor concentration was monitored continuous

and data logged using a newly purchased, calibrated AT

C16 PortaSenII with a hydrogen peroxide sensor (Analytical Technology, Collegeville, MD), which has a measurement range of 0 to 100 ppm HP vapor. The calib

was done on 9/6/07, directly before our experimenta

tests began; the manufacturer stated that the calibratio

had ±10% accuracy. Directly after our experimental test

were completed (1/30/08), the instrument was sent back

to the manufacturer to be re-calibrated. The instrumen

was reading 17% higher than it should have been—aga

within ±10% accuracy. Nevertheless, within the accur

of the calibration method, the instrument’s calibra

remained stable during our experimental test

After a constant HP vapor concentration had been

reached in the exposure chamber, test coupons were

inserted into the chamber through a vertically open

sliding door. When inserting test coupons, the door 

lifted only very slightly so that the HP vapor concentration would remain essentially const

Triethylene Glycol Vapor Tests. Tests in whic

influenza viruses deposited on stainless-steel coup

were exposed to TEG vapor were also done in the same

well-mixed 130-L cubical plexiglass chamber used for t

HP vapor tests. Greater care, however, was taken to seal 

the chamber, and dry air was not added to the chamber.

A shallow pool of 99% pure liquid TEG (VWR, Wes

Chester, PA) covered much of the floor area of the chamber. A beaker of water was placed within the chamber 

order to help maintain a reasonably constant RH. If th

beaker of water was not present, the RH in the chamber

would decrease over time because TEG is very hydroscopi

Temperature and RH were monitored and recorded every

30 s using a HOBO (Onset Computer Corp., Buzzards

Bay, MA). RH and temperature were also measured 

periodically with a hygrometer (Omega Engineer

and mercury thermometer. After equilibrium condit

had been achieved, test coupons were inserted into t

chamber by minimally opening a vertically sliding 

The coupons were inserted as quickly as possible so as 

minimize disruption of equilibrium c

The concentration of TEG vapor was not measured.

Because we allowed a large pool of nearly pure liqui

TEG located on the floor of the well-mixed exposure

chamber to reach equilibrium with the gas phase, the

was essentially saturated with TEG, and the partial pressure ( TEG p ) of TEG was approximately equal to its v

pressure ( o PTEG ). TEG vapor pressure can be calculated 

from the Antoine equation (NIST 2005

(1)

where vapor pressure is in bars and temperature (T) 

in degrees Kelvin. Based on Equation 1, the vapor pressure of TEG at 25°C is 0.00131 mm

, which is 

nearly perfect agreement with the value of 0.00132 mm

Hg at 25°C given by the EPA (2005a). The TEG mole

fraction ( TEG y ) in the gas phase can be calculated fro

Dalton’s law: 

(2)

where P is ambient pressure. In actuality, because li

TEG is so hydroscopic, the pool of TEG on the chambe

floor would tend to become diluted with water over ti

so that the mole fraction of TEG vapor would be somewhat less than 1.7 ppm. However, because the amount of

liquid TEG in the chamber was relatively large, dil

would not be expected to have a very significant effec

on the TEG vapor concentration

Tests of Natural Die-Off Rate. Our normal methodology for evaluating the loss of virus activity over

involved seeding 50 μL of influenza virus suspen

onto each of the coupons that were to be used during an

experimental test. All virus assays for an experimental te

were then done at the same time and, when possible, 

the same 96-well plate. Because growing and maintain

cells is somewhat of an art, performing all assays for a

experimental test at the same time is important in o

to get consistent results.

For tests to measure the natural die-off rate of influenza viruses, this methodology could not be used beca

the duration of the test was too long, so an alternativ

procedure was employed. In preparation for an experimental test to measure the natural die-off rate of influe

viruses, single-use packets of influenza virus suspen

1

Although Equation 1 was specified for a temperature range th

did not include 25°C, it predicted the same vapor pressure at 2

as was given by the EPA (2005a)

10 1.7 ppm 760

0.0013 6 = × =

= ≅

P

P

P

p

y

o

TEG TEG

TEG

1.299

3715 log 6.757 10 − = − T

Po

TEG 
were thawed, separated into 200-μL portions, and the

re-frozen at –80°C. At the start of a natural die-off test,

one of these 200-μL portions was thawed, and then each

of three stainless-steel coupons was seeded with 50 μ

of virus suspension prior to being exposed to amb

conditions in a small chamber without a decontamination agent present. After a predetermined amo

time, another 200-μL portion was thawed, and thre

additional coupons were seeded and then exposed t

ambient conditions. This procedure was repeated multiple times. At the end of the test, the virus res

each coupon was extracted using our standard method,

and each extraction was assayed at the same time and 

the same plate. 

Calculations 

The number of fluorescent focus units (FFU) per

volume of coupon rinsate is a measure of the quanti

of cultureable viruses present on the coupon. The rati

of the number of FFU per volume in the rinsate from a

exposed coupon (U) to the number from an unexposed 

coupon (Uo) is defined as the fraction of viruses rem

ing active ( f 

Uo

U f = (3)

For cultureable viruses, the number of log reduction

(n) is equal to the difference between the logarithm 

the initial FFU per volume and the logarithm of the fi

FFU per volume: 

n U U f o = log − log = −log (4)

where the logarithms are to the base 10. Thus, n = 4

corresponds to 4 log reductions, which is equivalent

0.01% of the viruses remaining active and 99.99% of 

viruses inactivated; that is, starting with 10,000 

the rinsate from an unexposed coupon, only one would

remain in the rinsate from the exposed coupo

Because three coupons were exposed and three were 

not exposed during a specific time period, the unexpo

and exposed coupons could not be separated into pairs

Therefore, the mean number of log reductions ( n ) was

calculated from Equation 5:

n = logUo − logU

where Uo log and logU are the means of the logarithms

of Uo and U, respectively. The standard deviation ( n s

corresponding to n can be calculated from the standard

deviation of log 

 ( Uo slog ) and the standard deviatio

of U ( U slog ):

2

log

2

log

2

n U o U s = s + s 

In figures in which the number of log reductio

( n ) is plotted versus time, the error bars correspond t

±1 standard deviation 

n

), as given by Equation 

Based on 109 FFU/mL for the influenza virus suspension in a single-use packet and 50% recovery of vi

from control slides, the theoretical limit of detec

in terms of the number of log reductions that could b

detected by the methods described above was calculated

to be 7.4. This calculation is based on the assumpt

that a single FFU detected from any of the three coupons

exposed at a specific test condition corresponds to t

limit of detectio

RESULTS

Hydrogen Peroxide Vapor Decontamination

The number of log reductions based on Equation 

versus exposure time for tests in which influenza vir

deposited on stainless-steel coupons were exposed a

approximately 25°C and 58-65% RH to relatively lo

concentrations of HP vapor is shown in Figure 1. In 

figure, error bars correspond to ± one standard deviation

based on Equation 6. Even at a HP vapor concentratio

as low as 10 ppm, about a two-log reduction was observed

after 2.5 min of exposure. The reduction, however, d

not increase as much as would be expected with increas

in either exposure time or HP vapor concentration.

a HP vapor concentration of 10 ppm and an exposure

time of 2.5 min are taken as the base, increasing expos

time by a factor of six or concentration by a factor 

nine added only an extra 1.6 and 1.3 logs of reduction

respectively. For 15 min of exposure time, the high

measured decontamination rate was 4.7 log reductio

at a HP vapor concentration of 90 ppm. An addition

test, not shown in Figure 1, in which influenza vi

were exposed at a HP vapor concentration of 57 ppm

for 60 min, resulted in a decontamination rate of 

log reductions

Triethylene Glycol Vapor Decontamination

The number of log reductions based on Equatio

5 as a function of exposure time for tests in wh

influenza viruses deposited on stainless-steel coup

were exposed to air saturated with TEG at 25-29°

and 45-55% RH is shown in Figure 2. Based on Equations 1 and 2, the concentration of TEG vapor in 

tests was equal to between 1.7 and 2.5 ppm. Error bars 

in Figure 2 correspond to ± one standard deviati

based on Equation 6. The number of log reduction

(n) versus exposure time (t) follows a linear relationship reasonably well. The relationship is given

following equatio

n = 1.31t (7)

where exposure time is in hours. Thus, the decontamination rate attributable to TEG vapor was 1.3 log redu

(5)

(6)
0

1

2

3

4

5

0 0.05 0.1 0.15 0.2 0.25

Exposure Time, hours

Number of Log Reductions

10 ppm

20 ppm

39 ppm

57 ppm

90 ppm

57 ppm

39 ppm

10 ppm

90 ppm

20 ppm

2.5 min 5 min 10 min 15 min

Figure 1. Surface decontamination of influenza viruses with hydrogen peroxide vapor 

n = 1.31 t

r

 2 = 0.78

-1

0

1

2

3

0 0.5 1 1.5 2

Exposure Time (t), hours

Number of Log Reductions (n)

Figure 2. Surface decontamination of influenza 

viruses with triethylene glycol saturated air 

per hour. Equation 7 is equivalent to Equation 8, 

equation for exponential decay of the fraction of viru

remaining active ( f 

 f = exp(–3.02t) (8)

Viability Tests at Ambient Conditions

For purposes of comparison with chemical decontamination tests, the natural die-off rate at ambient co

of influenza viruses deposited on stainless-steel coup

was measured. The number of log reductions versus tim

for two separate tests, each lasting a few days, is plotte

in Figure 3

Error bars in this figure correspond to ± one standar

deviation based on Equation 6. No decontamina

agent was used during these tests. Based on the data

points from both tests, the number of log reduction

(n) versus exposure time (t) follows a linear relations

given by Equation 

n = 0.0829t (9)

where exposure time is in hours. Thus, the natural dec

rate of influenza viruses was 0.083 log reductions p

hour, which is equivalent to a half-life of 3.6
DISCUSSION

Hydrogen Peroxide Vapor Decontamination

Test results on the decontamination of influenza viru

using HP vapor at concentrations less than 100 ppm (Fi

1) are somewhat surprising in that the number of l

reductions in active viruses versus exposure time is

non-linear; that is, the fraction of viruses remaining 

versus exposure time does not follow an exponential deca

curve. As the exposure time increases, the log reducti

rate decreases significantly; thus, as shown in Figure

the number of log reductions for the initial 2.5 mi

exposure is greater than the number of log reduction

from 2.5 to 15 min of exposure. This trend is true for a

HP vapor concentrations evaluated. For example, at a HP

vapor concentration of 10 ppm, the lowest concentration tested, the number of log reductions was 2.0, 3.

3.4, and 3.6 after 2.5, 5, 10, and 15 min of exposure,

respectively. Similarly, at a HP vapor concentration

90 ppm, the highest concentration tested, the numbe

of log reductions was 3.2, 4.5, and 4.7 after 2.5, 10, and

15 min of exposure, respectively

Another unexpected outcome of these tests was that, 

in the initial 2.5 min of exposure to 10 ppm HP va

the number of log reductions was equal to two, which 

a 99% virus reduction. If the number of log reductio

at 10 ppm HP vapor versus exposure time was linear

15 min of exposure would result in sterilization

log reductions). Instead, due to the nonlinearity of t

curves, after 15 min of exposure to 10 ppm HP vapor,

only 3.6 log reductions were measured. Nevertheless,

this is a significant reduction for such a low HP v

concentration. The Threshold Limit Value (TLV®) a

OSHA-permissible exposure limit (PEL)for occupati

HP vapor exposure is an eight-hour time-weighted aver

of 1 ppm (ACGIH 2008; OSHA 2008). This suggests

that 10 ppm of HP vapor is a relatively safe concentrati

over a short time period, although the TLV includes t

caveat that HP vapor is a “confirmed animal carcinog

with unknown relevance to humans.

Triethylene Glycol Vapor Decontamination

Dividing Equation 7 by Equation 9 indicate

TEG vapor increases the natural die-off rate of influen

viruses by a factor of 16. The decontamination rate f

air saturated with TEG vapor at 25-29°C, which w

measured to be 1.3 log reductions per hour, is considerably less than for HP vapor (Fig. 1), even at a concentration of 10 ppm. For example, for a 15-minute exp

period, the decontamination rate for TEG vapor w

0.33 log reductions, compared to 3.6 log reductions fo

HP vapor. Nevertheless, TEG vapor has some important

advantages. 

For surface decontamination using TEG vapor, ambient air or warmed air could easily be saturat

TEG prior to being introduced into an airplane c

Alternatively, micrometer-size TEG droplets, wh

evaporate rapidly, could be injected into the supply 

duct or directly into the cabin air. The standard met

for introducing TEG droplets into air for the purpose

air decontamination is through the use of a pressur

liquid (EPA 2005a), although a nebulizer could also 

used. If a pandemic were to occur, both surface and ai

decontamination could take place simultaneously, ev

while passengers were onboard

Although an objection could be raised due to th

potential health risk of using TEG vapor for air decontamination, this is likely an unwarranted concern

TEG is an odorless chemical of no known toxicity, 

exposure of people to TEG is already widespread. TE

vapor is used as a bacteriostat to kill odor-causing bact

for the purpose of air sanitation and deodorization. It

first registered for use in hospitals as an air disinfect

1947. Present application scenarios include spraying

inside offices, schools, hotels, lobbies, theaters, recept

rooms, sleeping rooms, bathrooms, and hospital room

(EPA 2005b). In addition, products containing 

packaged in aerosol cans and designed to be sprayed in

the air inside homes to control odors are sold in sto

everywhere (e.g., Oust® and Febreze®).

According to the U.S. Environmental Protecti

Agency (EPA 2005a), “the Agency has no risk concerns

for triethylene glycol with respect to human exposure

Based on a review of the available toxicology data, t

Agency has concluded that triethylene glycol is of ver

low toxicity by the oral, dermal, and inhalation rou

n = 0.0829 t

0

1

2

3

4

5

6

0 10 20 30 40 50 60 70 80

Exposure Time (t), hours

Number of Log Reductions (n)

7/14/2008

4/17/2007

best-fit line

(r = 0.82) 2

Figure 3. Natural die-off of influenza viruses on 

stainless-steel coupons at ambient conditions 
of exposure. The toxicology database is adequate t

characterize the hazard of triethylene glycol, and no dat

gaps have been identified. There are no indication

special sensitivity of infants or children resultin

exposure to triethylene glycol.” In addition, TEG h

no known deleterious effects on fabrics or other surface

(Lester, Kaye, Robertson & Dunklin 1950). Unlike H

vapor, TEG vapor is not an oxidizing agent. TEG inactivates viruses and bacteria because it is very hy

it condenses on bacteria- and virus-containing part

until the TEG concentration becomes sufficiently high

be germicidal (Puck, T.T. 1947a; Puck, T.T. 1947b

To demonstrate the effectiveness of simultaneou

surface and air decontamination, microbiological s

need to be conducted in a room-size chamber using bo

viruses and bacteria as challenges. Methods also need t

be developed to monitor TEG vapor concentration s

that TEG introduction can be precisely controll

It is reasonable to expect that the efficacy of TEG vapor

will increase as its concentration is increased. Howeve

25°C, the concentration of TEG in air cannot exceed 1

ppm because air is saturated at that concentration. T

only way of increasing the concentration is to incr

temperature. As shown in Table 1, which was calculate

from Equations 1 and 2, modest increases in temperatu

result in significant increases in TEG vapor concentration. Thus, further work investigating TEG va

decontaminating agent is warranted. Specifically, 

effectiveness of TEG vapor for surface decontamination at higher concentrations—that is, at tempera

greater than room temperature—should be determined.

In addition, the influence of RH on decontamina

effectiveness should also be evaluated

CONCLUSIONS

Our experiments show that HP vapor concentrations a

low as 10 ppm and TEG vapor concentrations of 2 ppm

can provide effective decontamination of a commer

airplane cabin. At these very low concentrations, t

potential for damage to the mechanical components o

avionics of the airplane would be expected to be mini

Although it has somewhat lower efficacy than 10 ppm HP

vapor, air saturated with TEG vapor at 25°C is probab

the better choice for decontamination of airplane cab

TEG is safer with regards to both personnel and airplan

and is easier to apply. If TEG vapor is determined to b

viable candidate for decontamination of airplane ca

optimizing temperature and RH would likely lead

greater efficacy.

REFERENCES

ACGIH (2008). TLVs and BEIs, American Conference

of Governmental Industrial Hygienists, Cincinnat

Burr, G.A., Van Gilder, T.J., Trout D.B., Wilcox, T.G.

and Driscoll, R.M. (1994). Health hazard evaluation, HETA 90-0355-2449. National Institute

Occupational Health and Safety.

De Benedictis, P., Beato, M.S., and Capua, I. (2007)

Inactivation of avian influenza viruses by che

agents and physical conditions: a review. Zoon

Public Health, 54(2), 51-68.

EPA (2005a). Reregistration eligibility decision for tri -

ethylene glycol, EPA739-R-05-002, United States

Environmental Protection Agency

EPA (2005b). Revised antimicrobials division’sreviewofthe

disciplinary sciences for issuance of the reregistration

eligibility decision (RED) document, EPA-HQOPP-2005-0002, United States Environmenta

Protection Agency

First, M.W., Rudnick, S.N., Banahan, K.F., Vincen

R.L., and Brickner, P.W. (2007). Fundamental

factors affecting upper room ultraviolet germici

irradiation—Part I. Experimental, J Occup Env

Hyg, 4(5), 321-31.

Hartshorn, K.L., White, M.R., Tecle, T., Tornoe, I.,

Sorensen, G.L., Crouch, E.C., and Holmskov, U. 

(2007). Reduced influenza viral neutralizing act

of natural human trimers of surfactant protein D

Respir Res, 8(1), 9-19.

Temperature, °C Concentration, ppm

20 1.1

25 1.7

30 2.8

35 4.4

40 6.9

Table 1. Triethylene Glycol Concentration in Air
Heckert, R.A., Best, M., Jordan, L.T., Dulac, G.C., Eddington, D.L., and Sterritt, W.G. (1997). Efficacy 

vaporized hydrogen peroxide against exotic an

viruses. Appl Environ Microbiol, 63(10), 3916-8

Lester, W., Kaye, S, Robertson, O.H., and Dunklin,

E.W. (1950). Factors of importance in the use o

triethylene glycol vapor for aerial disinfectio

J Pub Health, 40(7), 813-20.

Musher, D.M. (2003). How contagious are common

respiratory tract infections? N Engl J Med, 34

1256-66.

NIST (2005). Chemistry WebBook, National Institu

of Standards and Technology. <webbook.nist.gov

cgi/cbook.cgi?ID=C112276&Units=SI&Mas

#Thermo-Phase>.

Olsen, S.J., Chang, H.L., Cheung, T.Y.Y., Tang, A.F.Y., 

Fisk, T.L., Ooi, S.P.L., Kuo, H.W., Jiang, D.D.S

Chen, K.T., Lando, J., Hsu, K.H.H., Chen, T.J., 

and Dowell, S.F. (2003). Transmission of the sever

acute respiratory syndrome on aircraft. N Engl

Med, 349, 2416-22.

OSHA (2008). Limits for air contaminants, Stand

Number 29 CFR 1910.1000, Table Z-1, Electronic

Code of Federal Regulations. <http://ecfr.gpoaccess

gov/cgi/t/text/text-idx?c= ecfr&rgn=div5&vi

ext&node=29:6.1.1.1.1&idno=29#29:6.1.1.1.

.1.1.1>.

Puck, T.T. (1947a). The mechanism of aerial disinfection by glycols and other chemical agen

Demonstration that the germicidal action occ

through the agency of the vapor phase. J Exp Med, 

85, 729-39.

Puck, T.T. (1947b). The mechanism of aerial d

infection by glycols and other chemical agen

II. An analysis of the factors governing th

efficiency of chemical disinfection of the

Exp Med, 85, 741-57.

Robertson, O.H., Loosli, C.G., Puck, T.T., Bigg, E.

and Miller, B.F. (1941). The protection of mi

against infection with air-borne influenza

by means of propylene glycol vapor. Science, 94 

(2452), 612-3.

Robertson, O.H., Bigg, E., Miller, B.F., Baker, Z., an

Puck, T.T. (1941). Sterilization of air by cer

glycols employed as aerosols. Science, 93(2409), 

213-4. 

Robertson, O.H., Puck, T.T., Lemon, H.F., and Loosli,

C.G. (1943). The lethal effect of triethylene glycol

on air-borne bacteria and influenza virus. Scie

97(2510), 142.-4. 

Roy, C.J. and Milton, D.K. (2004). Airborne transmiss

of communicable infection—the elusive pathw

N Engl J Med, 350, 1710-2.

Rudnick, S.N. and First, M.W. (2007). Fundamenta

factors affecting upper-room ultraviolet germici

irradiation—Part II. predicting effectiveness. 

cup Environ Hyg, 4(5), 352-62. 

Schumb, W.C., Satterfield, C.N., and Wentworth, R.L.

(1955). Hydrogen peroxide, American Chemica

Society Monograph, series no. 128, Reinho

Publishing Corp., New Yor