Journal of Hospital Infection (1980) 1, 63-75

Sporicidal activity of glutaraldehydes and

hypochlorites and other factors influencing their
selection for the treatment of medical equipment
J. R. Babb, C. R. Bradley

and G. A. J. Ayliffe

Hospital Infection Research Laboratory, Dudley Road Hospital,

Birmingham B18 7QH
Summary:
Nine glutaraldehyde and 4 hypochlorite solutions were tested
for activity against approximately
10 spores of B. subtilis var. globigii in
suspension or dried on aluminium foil. All glutaraldehydes were effective
against the spores in suspensions in 3 h or less, but the acid glutaraldehydes
were much less effective than alkaline, particularly against the dried spores.
Although the acid glutaraldehyde preparations were more stable, they also
tended to be more corrosive. The buffered hypochlorite
solutions were
highly effective, killing the spores in less than 10 mix-r, but have the disadvantages of lack of stability and inactivation
by organic matter. The
buffered hypochlorites were relatively non-corrosive to metals, but damaged
rubber and the polyurethane coat of an endoscope after prolonged exposure.
All glutaraldehyde
preparations
passed the Kelsey-Sykes
capacity test
with and without yeast and using Pseudomonas aeruginosa as the test organism.
The hypochlorites failed the test when yeast was added.
Introduction

Glutaraldehyde is commonly used for the disinfection or sterilization of heat labile

medical equipment (Ross, 1966; Lowbury, Ayliffe, Geddes & Williams, 1975). In
recent years, particularly following the expiry of the patent on activated alkaline
glutaraldehyde (Cidex), a number of different glutaraldehydes have been introduced. Although most of these preparations contain 2 per cent glutaraldehyde, some
are alkaline and require an activator whilst others are acid and usually do not
require an activator
but have the advantage of greater stability. The sporicidal
activity of glutaraldehydes at room temperature tends to be rather slow, especially
when spores are dried onto surfaces, and results with the A.O.A.C. test indicate
that immersion for 10 hours or more is required for sterilization (A.O.A.C. 1975).
A more rapid sporicidal action has been claimed for buffered hypochlorite solutions
(Death & Coates, 1979).
In this study nine solutions of 2 per cent glutaraldehyde and four hypochlorite
preparations were examined for activity against Pseudomonas aeruginosa and spores
of B. subtilis var. globigii. Since these preparations may be used for treating expensive
medical equipment, some tests on corrosion and damage to materials were made.
The relevance of microbiological
and corrosion tests to practical problems of
disinfection and sterilization will also be discussed.
0105~6701/80/010063

@ 1980 Academic

13 $01.00/O

63

Press

Inc.

(London)

Limited

64

J. R. Babb et al.
Materials

and methods

Preparations of 2 per cent glutaraldehyde

1. Asep (Galen Ltd.) pH 7.6, liquid activated, green, stable for 14 days and contains corrosion inhibitor.
2. Acid glutaraldehyde, experimental formulation (not marketed) pH 5.5, stable
no activation required, colourless.
3. Cidex CX 250, (Arbrook Products Ltd.) pH 8.6, powder activated, green,
stable for 14 days and contains corrosion inhibitor.
4. Cidex Long Life (Arbrook Products Ltd.) pH 7.6, liquid activated, blue,
stable for 28 days, contains corrosion inhibitor, and a surfactant.
5. Alkaline glutaraldehyde,
experimental formulation
(not marketed) pH 8.6,
powder activated, blue, stable for 14 days, contains a corrosion inhibitor and
surfactant.
6. Clinicide (Bioscan) pH 6.2, powder activated, green, stable for 28 days, contains
a corrosion inhibitor and surfactant. (now marketed with an alkaline buffering
system)
7. 3M Instrument Disinfectant (3M UK Ltd.) pH 58, liquid activated, blue,
stable for 28 days, contains a surfactant.
8. Totacide (Tenneco Organics Ltd.) pH 7.5, liquid activated, green, stable for
28 days, contains a corrosion inhibitor and a surfactant.
9. Triocide (Vann Medical) pH 5.6-6.3, stable, no activation required, colourless,
contains a surfactant
All glutaraldehydes are supplied at in use concentrations with the exception of
Clinicide which is diluted l/10.
Preparations of hypochlorites
1. Anprosol (H. W. Andersen Products Ltd.) is a 0.2 per cent sodium hypochlorite
solution containing buffers and surface active ingredients, giving approximately
2000 parts/lo6 of available chlorine, pH 7.5. All the constituents are contained
in a triple compartment pouch which, when cut open, brings the disinfectant,
activators and corrosion inhibitors together. Additionally in each kit are provided
pouches of conditioning agent (detergent, wetting agents and corrosion inhibitors) for instrument preparation before immersion in Anprosol. The product
is supplied in a marked container suitable for dilution.
2. Sodium hypochlorite solution Sterite (Midland Direct Supplies Ltd.) made up
to give 1800 parts/lo6 available chlorine pH approximately 10.0.
3. Buffered sodium hypochlorite solution 250 parts/lo6 available chlorine pH 7.6
(Death & Coates, 1979).
4. Sodium hypochlorite solution Sterite made up to give 250 parts/lo6 available
chlorine pH approximately 9.0.
Titration of available chlorine
The available chlorine content of hypochlorites was assessed using the arsenite
method (Coates, 1977). 5 ml of hypochlorite was titrated with O-141 N or 0.0141 N

Sporicidal

activity

of glutaraldehydes

and hypochlorites

65

sodium arsenite and the end point detected by a failure to produce a blue stain on
starch iodide paper. The titration in ml, using 0.141 N, gives the available chlorine
concentration directly in g/l (i.e. 1 mEq to 1000 parts/106).
Measurement of pH
pH was measured before each microbiological
(Electronic Instruments Ltd). Glutaraldehyde
over a 2%day period.

test with a model 23A pH meter

solutions were measured weekly

Miuobiological
tests
Kelsey-Sykes capacity test. Tests were carried out as described by Kelsey &
Maurer (1974) for clean and dirty conditions. Glutaraldehydes
and hypochlorites were tested at in use concentrations using Ps. aeruginosa NCTC 6749 as
the test organism. Products were tested when freshly prepared, and the glutaraldehydes were also tested 14 days after activation.
Sporicidal activity : suspension test
Preparation of spore suspensions. Blood agar plates (Oxoid Columbia agar base
CM 331 + 7.5 per cent horse blood) were seeded with a freshly prepared
culture of Bacillus subtilis var. globigii (NCTC 10073) and incubated for 18 h at
37C. The resultant growth was removed with sterile cotton wool swabs and a
heavy aq. suspension prepared. The suspension was washed three times in
sterile distilled water, resuspended and heated to 56C and held for a period of
6 h. Spores were counted, using a surface dropping technique, and stored
overnight
at 4C to enable the spore challenge to be adjusted to approx.
lOr/ml. Sufficient spore suspension was prepared to test all the hypochlorites, and
glutaraldehydes over a 28 day period. Before each test, spores were heated and
counted by the method described.
Test method. 1 ml of spore suspension was added to 10 ml of the freshly prepared
hypochlorite
or glutaraldehyde
solution in a universal container (previously
rinsed in the disinfectant under test) and thoroughly mixed. 0.02 ml quantities of
this mixture were removed at specific time intervals-2,5,
10, 30 min and 1, 2, 3,4,
5, 6, 7 and 24 h, and added to each of a set of 5 recovery broths and thoroughly
mixed on a rotamixer. The disinfectant spore mixture was kept at room temperature (approximately 20C) throughout the period of the test and recovery broths
were incubated for at least 14 days and examined for growth of the test organism,
i.e. orange pellicle, granular and later turbid suspension. Doubtful tubes were
sub-cultured to confirm the test organism.
The test was repeated with the glutaraldehyde solutions at weekly intervals, i.e.
7, 14, 21 and 28 days after activation. The recovery medium was double strength
nutrient broth (Oxoid No. 2) with the addition of 10 per cent horse serum for the
glutaraldehyde tests and nutrient broth + 0.5 per cent sodium thiosulphate for the
hypochlorite tests.
Tests were carried out to establish that the recovery broth neutralized glutaraldehyde carried over during sampling and was not inhibitory to recovered test spores.

66

J. R. Babb et al.

To confirm the absence of surviving spores in the suspension test Bacillus

subtilis var. globigii spores were added to 10 ml of the freshly prepared disinfectants.
After 3 h the disinfectant/spore
suspension mixture was filtered (Millipore
0.45 PM), the membrane washed with 100 ml of serum nutrient broth and transferred to a blood agar plate. Membranes were transferred daily to fresh culture
plates for 14 days and examined for colonies of the test organism.
The suspension tests were repeated, with the addition of 2 ml of a 5 per cent
yeast suspension, on three of the glutaraldehydes (Asep, Cidex and Totacide)
showing good sporicidal activity and on the hypochlorites. The yeast, simulating
dirty conditions, was added before the spore suspension.
Sporicidal activity : carrier tests
Bacillus subtilis var. globigii spores (NCTC 10073) 106, deposited onto rolled aluminium foil from a 90 per cent methanol suspension and prepared by the method of
Beeby & Whitehouse (1965), obtained from Steriseal Ltd., Redditch, Worcs.,
were used as the test organism and carriers. These spores are marketed as control
organisms for the ethylene oxide sterilization process.
The spore strips were immersed in 10 ml of freshly prepared glutaraldehyde
or hypochlorite, mixed thoroughly on a rotamixer for 5 s and left for periods up
to 24 h at room temperature. At the same time intervals as in the suspension tests,
five strips were transferred with sterile wire hooks to each of a series of five recovery
broths. A set of five strips was immersed in a separate universal container for each
time interval.
The strips were shaken in the recovery broths for 5 s on a rotamixer, incubated
at 37C and examined for growth for periods up to 14 days. Tests were carried out
on freshly prepared hypochlorites, and glutaraldehydes at 0, 14 and 28 days after
activation. The pH and available chlorine content of the hypochlorites was
measured at the start of each test. Spores were recovered and counted from untreated foils by shaking them with glass beads in 10 ml of quarter strength Ringers
solution. Tenfold dilutions were made from these washings and plated out onto
the surface of nutrient agar plates using a dropping technique. Colonies were
counted after 24 h incubation at 37C.
Corrosion tests
Carbon steel and stainless steel scalpel blades were degreased, washed, dried and
immersed in beakers of freshly prepared glutaraldehyde or hypochlorite solutions
for periods up to 3 days. Each blade was placed in a separate container and supported
with nylon thread so that it was partially immersed in each of the disinfectants
under test. Untreated blades and blades immersed in tap water were used for
comparison with the treated blades.
In a second series of tests, various components of Olympus flexible fibreoptic
endoscopes, i.e. insertion tube sheath, nylon channels, guide wires, trumpet valves,
berylium/copper
and stainless steel protective coils and bending sections, were
immersed either in the 2 per cent glutaraldehyde solutions, 100 immersions for 3 h,
or in the hypochlorites, Anprosol, 100 immersions for 2 m in the conditioner

Sporicidal

activity

of glutaraldehydes

67

and hypochlorites

followed by 15 m in the hypochlorite, or for other hypochlorites 100 immersions of

20 m. Sections of rubber tubing and square pieces of anaesthetic equipment
(reservoir bags) were separately immersed. All items were washed, dried and
examined for signs of damage, e.g. rusting, tarnish, deposition, or loss of elasticity,
after each treatment or on a daily basis.
Results

Glutaraldehydes
All the glutaraldehydes passed the Kelsey & Sykes test at in use dilution under
clean and dirty conditions. No growth was obtained in any of the recovery broths.
Tests with added spores showed that adequate neutralization of the disinfectant
had occurred.
Table I. Sporicidal activity of 2 per cent activated and 2 per cent stable
glutaraldehydes : Suspension test (inoculum l-4-7.4 x 10 spores of B.
subtilis var. globigii)

Agent

RecomNo. of tubes showing growth after 14 days

incubation
mended
postexposure time
activation Day of
life (days)
test
10min 30min
lh
2h
3h 4h
pH

(a) Sporicidal

activity of 2 ner cent activated glutaraldehvdes

0
315
Asep
::
515
14
:;:
0
515
515
0
0
515
275
Cidex
515
5):
14
:;:
515
515
515
575
575
Alkaline
14
1:
515
glutaraldehyde
28
515
515
5/5
0
0
0
315
Cidex Long Life
515
28
;;:
00
i
515
0
5/j
Clinicide
:
:
515
28
ii
515
515
515
0
0
515
275
28
14
Totacide
515
45:
28
415
515
415
515
3M
$5
28
104
515
28
315
515

00

(b) Sporicidal activity of 2 per cent stable glutaraldehydes

Acid glutaraldehyde
15/15 15/15
Triocide
15/15 15/15

10/15
s/15

:
0
0

0
0
0

575
t

175
0

515
ii

41:
0

:
0
0
:

0077:;
0

7.2

i
0

t:;:
7.6

0
:

;:;
7.8

i
0

5:;
7.4

:
0

2:;
6.0

:
0
0
0
0
0

00
0
0

77:;
7.1
5.8

5.8
5.9

0
0

0
0

5.5
5.6

4115
s/15

Table I(a) shows the sporicidal activity of the activated glutaraldehydes, and I(b)
the stable glutaraldehydes, when the test organism (Bacillus subtilis uar. globigii)

J. R. Babb et al.

68

was added in suspension. Table I(b) also indicates the pooled results of sporicidal
activity of the stable products on three occasions over a 1 month period. All
products killed 10 spores within a three hour period, providing the post-activation
period, recommended by the manufacturer, was not exceeded, i.e. 14 or 28 days.
This apparent kill was substantiated by failure of the spores to grow on the surface
of the cultured membrane filter after daily transfers. Many of the products (Asep,
Cidex, Cidex Long Life, Totacide, and Triocide) killed the spores in less than
1 h.
Table

II. Sporicidal

No. of tubes showing growth after 14 days incubation

Time of exposure
10 min
30 min
lh
2h
3h

Agent
Asep
Cidex
Totacide
Spore

activity of 2 per cent glutaraldehydes : Suspension

test with added yeast

challenge

515
515

515
515

515
515

515

515

515

575

3.0 x 10

The pH of the alkaline formulations (Cidex, Asep, and Totacide) dropped

over the 2%day test period and this fall appeared to be associated with their
sporicidal activity. The pH of the acid formulations, including the activated acid
products (3M, and Clinicide), remained constant over the test period although
the sporicidal activity varied considerably. Table II shows the sporicidal activity of
Asep, Cidex, and Totacide when organic material (yeast) was present in the
suspension. 10 spores were killed within 3 h. These three products were chosen
as they did not appear to damage or corrode immersed materials in preliminary
tests and were reasonably effective in the surface spore tests.
The survival of Bacillus subtilis var. globigii dried on rolled aluminium foil and
immersed in the 2 per cent glutaraldehydes is shown in Table III(a), (b). The
differences between the products are more clearly defined than in the suspension
tests and do not appear to be related to the presence of surfactants. The acid
formulations (acid glutaraldehyde and 3M) required 7 or more hours to kill the
spores. All other products killed the spores in under 3 h although the same loss of
activity was noticed with the 14 day products when this period was extended. The
mean number of spores recovered per foil was 1.9 x 10 (range 8.0 x 10G2.3 x 107) which is approximately 1 log higher than stated by the manufacturer.
Hypochlorites
The sporicidal activity of freshly prepared hypochlorites (Table IV suspension
test, Table V surface test), is more rapid than the glutaraldehydes, especially
when buffered to a pH of approximately 7.6. Anprosol (1800 parts/lo6 of available
chlorine, pH 7.5) and buffered sodium hypochlorite (pH 7.6, loo-250 parts/l06 of
available chlorine), as described by Death & Coates (1979) killed 10 spores in under

0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0

I4
L-S
8-S
E*L
S-L
E*L
P-9
t-9
2.9
E*L
C-L
9.L
8-L
1.8
8.8
9-L
1.8
9.8
0.L
Z-L
9-L

Hd4Pz

0
0

E.9
s*s

uopeqnsu!

Sk3p

Y9

UE

Yb

92

s/s

sls
SIS

0
SI/SI

8U!MOlfS

amy amsodxa

3:

S/S

ml

s/s

S/l
s/s

0
sr/sr

+I Jayr3 qJMOI8

9s

Sk

Sk

YL

s/z

0
0
0

0
srlsr

0
srlsr

0
srlor

SaqW

YI

si:

sls

srlor
srls1

30 ON

01

was

30 A=a

8Z

S/S

U1l.U

82

S/S

O&

ii
0

(shnep)a3tt
uoga.+a-Jsod
papuaunuoDa8

82

87:

82

3uaZv

Saw,

xaP!S

aP!~~oLTAL,

IN&,

(q)

aP!~o!JLLL,
apLqap~s.mn~% pf3hT
saplCqappmz)nl% aIqels luau lad z 30 LJ!A~X Iapf+odS

sls
sls
sls

SIISI
SIISI

U1U.I

sls

s/s
sls

SIISI
SIISI

J. R. Babb et al.

70

Table IV. Spmicidal activity

of hypochlorite solutions : Suspension test

No. of tubes showing growth after

14 days incubation
Available
Time of exposure
chlorine
2
30 1 h 2 h
(parts 106) min nZn rZn min

Agent

Anprosol
pH 7-S
Hypochlorite
pH approximately

10

Buffered hypochlorite
pH 7.6

Hypochlorite
pH approximately

;;;;

w:

215
0

1800
1800
2100
1200
1700
1800
loo
100
150
150
250
100
loo
150
150
250

O/S
O/S
o/5
s/5
515
515
s/5
415
o/5
o/5
O/5
515
s/5
515
315
s/5

0
0
0
5/s
515
515
315
0
0
0
0
515
515
s/5
l/5
515

0
:
315
s/5
515
415
8
0
0
515
515
5/s
215
515

00
0
0
415
5/S
0
0
00

:
0
0
0
0
0
0
0

ii
0
0

0
l/5
415
315
0
515

ii
0
0
0
0
0

Spore count
w
3.0 x 10
5.4
8.8
4.4
3.1
5.4
3-o
4.4
4.4
3.1
8-8
5.4
1.9
3-l
4-4
5.4
8-8
3.0

:
a0
:
0
:
!
0

x
x
x
x
x
x
x
x
x
x
x
x
x
x
x
x

106
lo6
106
10
10
106
106
106
106
10
10
106
106
10
106
10

10 min and often in under 2 min. Unbuffered preparations were less effective, and
times required to kill 10 spores were comparable with those of the better glutaraldehydes.
Table V. Sporidal

Agent

activity

of hypochlorite solutions : Surface test

Available
chlorine
(parts 103

No. of tubes showing growth after 14 days

incubation
Time of exposure
2min
5 min
10 min 30 min
1h

Anprosol
pH 7.5

1800

015

Hypochlorite

1800

515

515

515

250

015

250

515

515

515

315

pH approximately 10
Buffered hypochlorite
pH 7.6
Hypochlorite
pH approximately 9
Mean spore count per strip 1.9

10

In the presence of yeast, all hypochlorites failed to kill spores even when the
exposure period was extended to 24 h. These failures were also shown in the
capacity test of Kelsey & Sykes. The activity of the hypochlorites diminishes fairly

Sporicidal

activity

of glutaraldehydes

and hypochlorites

71

rapidly after preparation, particularly with lower dilutions, as does the available
chlorine concentration, e.g. Anprosol showed a fall from 1800 to 1200 parts/l06
in 24 h, and buffered hypochlorites from 250 to 130 parts/lOs.

corrosiontests
The alkaline glutaraldehydes Asep, Cidex, Cidex Long Life and Totacide did
not appear to cause damage to carbon steel or stainless steel scalpel blades when
immersed for periods up to 3 days. Triocide on the other hand showed some
deterioration of carbon steel (rust and tarnish) after a short period (3 h) of immersion and the stainless steel blade, separately immersed, was rusted within 3
days. Immersion in an acid glutaraldehyde, e.g. Clinicide and 3M also produced
damage to carbon steel blades although there was no damage to stainless steel.
This effect was minimal in 3 h, although considerable at 3 days.
The interpretation of tests on other materials was more difficult. The buffered
alkaline formulations Cidex, Cidex Long Life and Totacide caused no visible
damage to collections of endoscope components after 100 immersions of 3 h. The
acid formulations
Clinicide,
Triocide and 3M, became cloudy and heavy
deposits formed making visual examination of components difficult. Some damage
was certainly caused to collections of dissimilar metals particularly copper alloys.
Anprosol (pH 7*5), and buffered hypochlorites (pH 7.6) caused little damage to
immersed scalpel blades and metal endoscope components although non-buffered
hypochlorite, pH 9-10, caused severe corrosion of carbon steel within a few hours.
including the
Two problems were, however, noticed with the hypochlorites,
buffered formulations, especially at higher concentrations. A thick, white deposit
built up on the insertion tube of the Olympus flexible fibreoptic endoscopes during
100 immersions and although this could be scraped or wiped off, the polyurethane
coating appeared to be destroyed. Sections of rubber reservoir bags and tubing
were also damaged. The rubber hardened, lost its resilience, and cracks appeared
on the surface when stretched.
Discussion
A reproducible

quantitative suspension test measuring the log reduction in numbers

of organisms would have been preferable to the technique used (Reybrouck &
Werner, 1977). However, in view of the large number of tests, this was not possible
and a semi-quantitative method was used to assess sporicidal activity. The results
provide a useful comparison between the agents tested but do not necessarily
indicate exposure times required for sterilization in practice.
Spores vary in their resistance to disinfectants, depending on their method of
preparation, but the spores of B. subtilis var. globigii, as used in these tests, are
likely to be more resistant to glutaraldehydes and hypochlorites than pathogenic
species. The numbers used were also considerably higher than would be found on a
cleaned instrument. The preparation of a reliable spore suspension is often difficult
and the commercial spore strips proved to be consistent in both numbers of spores
and in response to the chemical agents. Glutaraldehyde is not easily neutralized but
dilution in broth containing 10 per cent serum was found to be as reliable as other

72

J. R. Babb et al.

potential neutralizers and is not toxic to damaged organisms (Russell, Ahonkhai &
Rogers, 1979). Attempts to recover damaged spores using a membrane filter
technique also failed.
All the glutaraldehyde preparations passed the Kelsey-Sykes capacity test, and
killed spores in suspension in 3 h. The alkaline glutaraldehydes were generally
more active than the acid preparations some of which required long periods to kill
spores dried onto metal carriers. The presence of a surfactant did not appear to
improve the effect. Although the A.O.A.C. test suggests a 10 h exposure is necessary to achieve a complete sporicidal effect (A.O.A.C. 1975), 3 h should be sufficient
for practical purposes, particularly as spores are infrequent on clean medical equipment (Spaulding, 1978). Th e immersion, at 20C of five or more commercial
spore suspensions dried on metal foil could provide a useful laboratory screening
test for glutaraldehydes. This test should be done on activated glutaraldehydes,
stored at 20C on their expiry date, i.e. at 14 or 28 days after activation.
Opinions on the tuberculocidal activity of 2 per cent glutaraldehyde at room
temperature vary but the immersion times required are considerably shorter than
for spores and 20 m is generally considered adequate (Bergan & Lystad, 1971,
Miner et al., 1977).
A possible advantage of using acid glutaraldehydes is stability. When heated to
55-60C their sporicidal activity is greatly enhanced (Sierra & Boucher, 1971),
and in our tests lo6 spores were killed in less than 5 min at this temperature. The
differences between the sporicidal activity of the alkaline and acid glutaraldehydes
diminishes with a corresponding increase in temperature. However, special equipment would be required for this treatment and may have to include equipment for
the removal of toxic vapours. An acid solution of 2 per cent glutaraldehyde (pH
3-4) is stable for several years. However, when alkalinized to produce the optimum
sporicidal activity the corresponding loss in active aldehyde groups, and hence
stability, limits its use to 14 days. By stabilizing the pH to approximately 7.6 the
period of activity of alkaline glutaraldehydes can be extended, with no loss in
sporicidal activity, to 28 days in situations of low dilution (Miner et al., 1977).
This is supported by our findings although the interpretation of these results in the
practical situation should be treated with some caution.
In this study, the acid glutaraldehydes were stable and the activated glutaraldehydes showed little loss in activity over 14 days. However, the length of time a
disinfectant is repeatedly used depends on the degree of dilution, the amount of
organic and other materials added to it, and storage conditions as well as its stability.
Although 2 per cent glutaraldehyde is not readily inactivated by organic materials,
repeated use of the same solution over long periods is not good practice. Disinfectant solutions should preferably be discarded after each use, but this practice
may be too expensive with glutaraldehyde. If used infrequently and for clean eyuipment only, 7-14 days use would seem to be a useful compromise, but the decision
should be made by the microbiologist
depending on the particular use of the
solution.
The interpretation of the corrosion tests presented some problems. The acid
formulations usually caused greater damage to immersed metals and as their spori-

Sporicidal

activity

of glutaraldehydes

and hypochlorites

73

tidal activity is less rapid, longer immersion times may be required. Carbon steel is
rarely incorporated in expensive equipment and corrosion of this alone is of doubtful
practical significance. Nevertheless, it seems reasonable to choose a product which
contains a corrosion inhibitor for disinfection or sterilization of expensive equipment. There is always the possibility of accidental long exposures, e.g. over a weekend. It is obviously less important when disinfecting inexpensive equipment or
using short exposure times, but it is advisable to carry out in-use tests or to consult
the instrument manufacturer before changing from one preparation to another.
The hypochlorite solutions have several potential advantages. They kill spores
rapidly and instruments could be sterilized rather than disinfected between
patients during busy endoscopy sessions. Hypochlorites are cheap and could be
discarded after each use or after an operating session. However, the cost of essential
buffers and corrosion inhibitors is likely to increase considerably this cost. These
solutions have been used for disinfecting infant feeding bottles for many years and
dilute solutions are relatively non-toxic. The main disadvantages of hypochlorites
are inactivation by organic material, instability at low concentration, and possible
damage to instrument components. Two of the buffered preparations examined,
pH 7-S-7.6 (Anprosol and buffered hypochlorite),
showed surprisingly
little
damage to metals but did cause some damage to the outer layer of the insertion
tube of an Olympus flexible fibre-optic endoscope on prolonged exposure, and to
rubber items, e.g. anaesthetic equipment. We have also received complaints that
endoscope eye piece mounts are likely to bleach during prolonged or frequent
immersions in hypochlorite and that surfactants and conditioning agents, used
prior to or during disinfection, remove essential lubricants and these should be
replaced. It could be argued that only short exposures are required, but accidental
immersion for long periods is always possible particularly if staff are familiarised
with the use of glutaraldehydes. However, if the risk is known, users tend to be
more careful, e.g. alcoholic solutions of chlorhexidine
are sometimes used for
disinfection of cystoscopes, and exposure for longer than a few minutes may damage
the lens mounting.
Heat is the most reliable method of disinfection or sterilization, and the use of
chemical solutions of disinfectants or sterilants should rarely be required in
hospitals, but autoclaving at high temperature will damage most endoscopes and
destroy flexible fibrescopes. Low temperature steam at 73C for 10 min is perhaps
the most suitable method for disinfecting cystoscopes between patients, and low
temperature steam with formaldehyde for longer periods for sterilizing laparoscopes
and arthroscopes (Alder, Gingell & Mitchell, 1971). Low temperature steam is also
particularly suitable for the disinfection of respiratory equipment, where hypochlorites cannot be used as rubber is damaged, and inadequate removal of glutaraldehyde is always a potential hazard to the patient. However, low temperature
steam machines are not always available and reliable low temperature steam and
formaldehyde machines are still being developed (Cripps, Deverill & Ayliffe, 1976).
Low temperature steam, especially with formaldehyde, will damage most of the
flexible fibre-optic endoscopes in use at the present time. Chemical disinfection is
therefore required and 2 per cent glutaraldehyde is commonly used despite the

74

J. R. Babb et al.

problems of penetration of narrow tubing and valves and the necessity of thorough
rinsing. Treatment of flexible fibre-optic endoscopes with glutaraldehyde for a
short period, between use on different patients, is a useful compromise (Axon,
Phillips, Cotton & Avery, 1974; Ayliffe & Deverill, 1979; Noy, Harrison, Holmes
& Cockel, 1980). Cleaning according to the manufacturers instructions is more
effective, but takes up to 20 min. Although exposure to glutaraldehydes for 3060 min should be adequate following use of an endoscope in a patient with tuberculosis, hepatitis or salmonellosis, treatment of the cleaned instrument with ethylene
oxide will further increase the margin of safety.
Treatment with glutaraldehyde for lo-20 min will disinfect but not sterilize, yet
laparoscopes and arthroscopes are often treated for this short time. Despite
inadequate sterilization infection appears to be rare, presumably because there are
few potentially pathogenic spores on the cleaned endoscope. The use of hypochlorites could considerably increase the reliability of this procedure and further
studies are in progress. Endoscopes are expensive, and often complex, and are
difficult to clean, disinfect, or sterilize. It is hoped that manufacturers will produce
instruments that will withstand autoclaving at high temperatures without shortening the life of the instrument.
We wish to thank KeyMed for supplying the Olympus endoscope components used
for disinfectant immersion studies and the disinfectant manufacturers for the products
tested.

A.O.A.C. Official methods of analysis of the Association of Official Analytical Chemists.

Journal of Association of OjjWul and Analytical Chemists. 12th edition. 57-71 (1975).
Alder, V. G., Gingell, J. L. & Mitchell, J. P. Disinfection of cystoscopes by subatmospheric
steam and steam and formaldehyde at 80C. British MedicalJournal iii: 677-680 (1971).
Axon, A. T. R., Phillips, I., Cotton, P. B. & Avery, S. A. Disinfection of gastrointestinal
fibre endoscopes. Lancet i: 656-658 (1974).
Ayliffe, G. A. J. & Deverill, C. E. A. Decontamination
of gastroscopes. Health and Social
ServicesJournal May: 538-540 (1979).
Beeby, M. M. & Whitehouse, C. E. A bacterial spore test piece for the control of ethylene
oxide sterilization. Journal of Applied Bacteriology 28: 349-360 (1965).
Bergan, T. & Lystad, A. Antitubercular
action of disinfectants. Journal of Applied Bacteriology 34: (4): 751-7.56 (1971).
Coates, D. Kelsey-Sykes capacity test: origin, evolution and current status. Pharmaceutical
&urnizl219:
402-403 (1977).
Cripps, N., Deverill, C. E. A. & Ayliffe, G. A. J. Problems with low-temperature steam and
formaldehyde sterilizers. Hospital Engineering AuglSept International Federation Issue
no. 19 (1976).
Death, Janet E. & Coates, D. Effect of pH on sporicidal and microbicidal activity of buffered
mixtures of alcohol and sodium hypochlorite. Journal of Clinical Pathology 32: 148-153
(1979).
Kelsey, J. C. & Maurer, I. M. An improved Kelsey-Sykes test for disinfectants. Pharmaceutical Journal 213: 528-530 (1974).
Lowbury, E. J. L., Ayliffe, G. A. J., Geddes, A. M. & Williams, J. D. In Control of Hospital
Infection : A Practical Handbook Chapman & Hall: London (1975).
Miner, N. A., McDowell, J. W., Willcockson, G. W., Bruckner, N. I., Stark, R. L. &
Whitmore, E. J. Antimicrobral and other properties of a new stabilised alkahne glutaraldehyde disinfectant/sterilizer.
American Journal of Hospital Pharmacy 34: 376-382
(1977).

Sporicidal

activity

of glutaraldehydes

and hypochlorites

75

Noy, M. F., Harrison Lynne, Holmes, G. K. T. & Cockel, R. The significance of bacterial
contamination of fihreoptic endoscopes. Journal of Hospital Infection 1: 53-61 (1980).
Reyhrouck, G. & Werner, H. P. Ausarbeitung eines neuen quantitaven in-vitro-tests fur die
bakteriologische Prufung chemischer Desinfektionsmittel.
Zentralbatt fur Bakteriologie,
Parasitenkunde, Infektionskrankheiten und Hygiene, I. Abteilung originale Reihe B 165:
126-137 (1977).
Ross, P. W. A new disinfectant. Journal of Clinical Pathology 19: 318-320 (1966).
Russell, A. D., Ijeoma Ahonkhai & Rogers, D. T. A review: microbiological applications of
the inactivation
of antibiotics and other antimicrobial
agents. Journal of Applied
Bacteriology 46: 207-245 (1979).
Sierra, G. & Boucher, R. M. G. Ultrasonic synergistic effects in liquid-phase chemical
sterilization. Applied Microbiology 22: 160-164 (1971).
Microbiological
Spaulding, E. H. Fibreoptic endoscopes: disinfection or sterilization.
aspects of the dilemma. Hospital Infection Control 5: 35-39 (1978).