Anaesthesia, 1993, Volume 48, pages 30-32
Growth of micro-organisms in solutions of intravenous anaesthetic
agents
C. B . BERRY, T. GILLESPIE, J . HOOD
AND
N. B. SCOTT
Summary
This contrnlled bench study compared the growth of' bucteriu and yeast in solutions of commonly used intravenous anaesthetic
agents at r ~ o mand body temperutures. Drugs presented in aqueous emulsions support both hucterial und.funga1 growth. Five out
of the s i s solutions supported the growth of yeast. The one exception was chlormethiazole. which is both ,fungicidal und
bactericidul. Solutions ?f' drugs which will support growth of micro-organisms should be used with great cure and they should not
be stored or infused over prolonged periods.
Key words
Anaesthetics, intravenous.
Bacteria; growth rates.
Propofol is an intravenous anaesthetic agent which is used
for the induction and maintenance of anaesthesia. It is also
an effective sedative both for short term endoscopic procedures and for longer term sedation of patients in intensive care units (ICU). It can be administered either by bolus
injection or by continuous infusion.
Since it is a highly lipid soluble drug, propofol is
presented as an aqueous and isotonic emulsion containing
glycerol, purified egg phosphatide, sodium hydroxide, soya
bean oil and water, similar to a lipid emulsion used in
parenteral feeding. Anecdotal reports have suggested that
propofol supports bacterial growth [ I ] and the manufacturer's recommendations are that the unused contents of
open ampoules should be discarded. I t is, however, not
uncommon for large volume infusions to be administered
over several hours in both ICU and operating theatres.
The purpose of this laboratory study was to determine
the potential for growth of micro-organisms in propofol
solutions at room and body temperatures. In addition, we
have cornpared propofol in this respect with other
commonly used solutions of intravenous anaesthetic and
sedative agents.
Methods
Six test solutions and two controls were inoculated with
thrce standardiscd cultures of micro-organisms and colon-
isation of each solution was measured at 6, 12, and 24 h at
both 20°C and 37°C.
The pharmaceutical preparations tested were: Hypnovel
(Roche), midazolam hydrochloride, 2 mg.ml-' in aqueous
solution, pH 3.3; Diprivan (ICI), propofol, 10 mg.ml-I in
aqueous isotonic emulsion containing glycerol, purified egg
phosphatide, sodium hydroxide, soyabean oil and water.
pH 7.1-7.2; lntraval sodium (Rhone-Poulenc Rorer), thiopentone sodium 25 mg.ml-' in aqueous solution with
sodium carbonate (60 mg per gram of thopentone),
pH 10.5; Heminevrin (Astra), chlormethiazole edisylate
8 mg.ml-' in aqueous solution containing 4 % dextrose and
sodium hydroxide to adjust pH to 6-7.5; Diazemuls
(Dumex), diazepam 5 mg.ml-' in isotonic aqueous emulsion containing glycerol, purified egg phosphatide, sodium
hydroxide, soyabean oil and water, pH 8; lntrahpid 10%
(Kabivitrum), oil in water emulsion containing fractionated
soyabean oil, fractionated egg phosphatide. glycerol and
water, pH 7.
The control solutions were: sodium chloride 0.9%;
Robertson's cooked meat broth. The following organisms
were used to inoculate the test solutions: Pseudomonu.s
ueruginosa (NCTC 10662); Stuphj*lococcus sp. (coagulasc
negative); clinical isolate (IN 25 I ); Cundicfu ulhicms;
clinical isolate (IN 252).
Standardised suspensions of organisms were produced
by innoculating broth (Isosensitest broth for bacteria and
C.B. Berry. MB, BS. FRCAnaes. Registrar in Anaesthesia, T. Gillespie. BSc, MB, ChB, Senior House Oficer in
Microbiology, J. Hood. BSc, PhD, MB, MRCP. MRCPath. Consultant in Microbiology, N.B. Scott. MR, ChB, FRCS.
FFARCS(I), Lecturer in Anaesthesia. Glasgow Royal Infirmary. Castle Street, Glasgow G 4 OSF.
Correspondence to Dr Scott please.
Accepted 26 June 1992.
0003-2409/93/O 10030f03 %08.00/0
@ 1993 The Association of Anaesthetists of Gt Britain and Ireland
30
31
Micro-organisms in anaesthetic agents
I2l
'r
P
10
'I1
:\
I
Time(h)
.,
Fig. 1. Growth of Pseudomonus at 20°C.
Hypnovel; 0 ,
Diprivan; Intraval; 0, Heminevrin; A, Diazemuls; A. Intralipid;
0 , saline; 0.broth.
*,
brain-heart infusion for yeast) with each organism and
incubating overnight at 37°C. Test solutions were then
inoculated with known amounts of the suspensions to give
an approximate initial concentration of 10) colony forming
units per millilitre (cfu.ml-I). Samples were taken from
each culture at 6, 12 and 24 h and plated out on Columbia
blood agar for a further 24 h incubation. The number of
colony forming units was determined by the Miles and
Misra's dilution technique [2].
The inoculated solutions were incubated for 7 days at
4°C and tested for viable organisms.
Time ( h )
24
Fig. 2. Growth of Pseudomonus at 37°C. For explanation of
symbols, see Figure I legend.
10 r-
99 t
8c
7-
It
0
\
6
Results
All counts were recorded as logarithm cfu.ml-' for ease of
data interpretation and graphical representation. A count
of 0 cfu.ml-l was only recorded if viable organisms could
not be grown from a sample after further incubation in
nutrient broth. The count in each sample was approximately 10' cfu.mI-l after initial inoculation at time zero.
The positive control (broth culture) supported growth as
expected at both 20'C and 37 C (Figs 1-6).
Saline 0.9% solution allowed bacterial counts to be
sustained at a static level for up to 12 h after which counts
decreased. Counts of the yeasts increased in saline 0.9% at
a rate less than that in nutrient broth (Figs 5 and 6).
Heminevrin, Intraval and Hypnovel all demonstrated
bactericidal activity, with no growth of either the
Stuphj~lococ'cus or the Pseudonionas at 6 h or beyond
(Figs 1-4).
Cundidu counts were seen to increase in all solutions
other than Heminevrin from 6 h after inoculation at both
22'C and 37°C (Figs 5 and 6). There was little growth of
yeasts in aqueous solutions at 12 h but, by 24 h. a 10' fold
increase was recorded in lipid emulsion containing solutions and saline. Growth in lntraval and Hypnovel was of a
lower magnitude. No viable Cundidu were found in the
Herninevrin solution at 6 h or beyond.
Diprivan, Diazcmuls and lntralipid solutions all
sustained growth of bacteria and yeast at both 20'C and
37'C. Staphylococci grew more rapidly in lipid emulsion
solutions than broth at 20 C with 10-fold increases being
recorded at 6 h and lo4 fold increases by 24 h (Fig. 3). At
37' C the growth of Sruph.dococci in these solutions was
similar but was more consistent with the rate of growth in
broth (Fig. 4).
12
6
0
-
0
12
24
Time ( h )
Fig. 3. Growth of SIuphylococcus at 20°C. For explanation of
symbols, see Figure I legend.
'r
7-
-
6-
I
0
\24
6
Time ( h )
Fig. 4. Growth of Siuphglococcus at 37°C. For explanation of
symbols. see Figure I legend.
The growth of Pseudomoms in broth was very similar to
that in lipid emulsion preparations. At 20°C a 10-fold
increase was recorded at 6 h and by 24 h, a 105-fold
increase was recorded (Fig. I). At 37"C, the rate of growth
was increased with a 103-fold increase at 6 h and a 107-fold
increase at 24 h (Fig. 2).
Viable organisms were obtained from Intralipid,
Diprivan, Diazemuls and nutrient broth incubated at 4°C
up to 7 days postinoculation. No viable organisms were
isolated from the other preparations after 7 days at 4°C.
32
C.B. Berry et al.
111
r
Time ( h )
Fig. 5. Growth of Candida at 20°C. For explanation of symbols,
see Figure 1 legend.
Time ( h )
Fig. 6. Growth of Candida at 37°C. For explanation of symbols,
see Figure I legend.
Discussion
Our results have shown conclusively that both Diprivan
and Diazemuls provide a fertile culture medium for the
bacteria and yeast tested. This is true both at room
temperature and body temperature. Recently there have
been anecdotal reports of postsurgical infections related to
the use of propofol [ I , 31. No data have been published
comparing propofol in this respect with other commonly
used anaesthetic agents.
The manufacturer's recommendations emphasise that
propofol will support the growth of micro-organisms and
that the drug should be used immediately once an ampoule
or vial has been opened. However, infusions of propofol
may last for several hours either in the operating theatre or
ICU. It is usual procedure in this hospital for infusions to
be prepared at the site of use either by doctors or nurses.
The risk of colonisation might be minimised if small
volume infusions were made up complete with giving sets,
under sterile conditions, by pharmacy staff [4].However,
the use of custom made infusion vials may still allow
contamination of the solution with micro-organisms at the
time of insertion of the giving set, despite an aseptic
technique, a condition which may be difficult to guarantee
in a busy clinical environment.
Multi-use vials of thiopentone have been employed for
many years and, interestingly, our results show that this
solution is bactericidal. The yeast may survive in it at 20"C,
but without replication; however, at 37°C survival is not
sustained past 24 h. It is probable that the bactericidal
effect is a consequence of the relatively high alkalinity of
this particular thiopentone solution. Evidence for a pHrelated mechanism for the bactericidal action of thiopentone and midazolam has been supported by further experiments carried out by us (unpublished data). However, the
mechanism of the bactericidal and fungicidal action of
chlormethiazole is less clear and is at present being
investigated.
This study clearly shows the importance of a scrupulous
aseptic technique in the preparation of infusions of intralipid-based anaesthetic and sedative agents in clinical practice. It is also prudent to ensure that individual infusions,
once prepared, should not be stored or infused over
prolonged periods, since this may allow small numbers of
contaminating bacteria to multiply into a significant
bacterial challenge. Therefore if these solutions are not
used correctly, they have the potential for increasing the
problem of nosocomial infections, especially in individuals
with increased susceptibility to infection. This is particularly relevant to patients in ICU.
However, we would wish to stress that, although some
agents used in this study appear to inhibit the growth of
bacterial and fungi, meticulous attention should still be
paid to the details of preparation and administration.
Many intravenous drugs have the potential, under the right
conditions, to act as vehicles of infection. It is our duty as
medical practitioners to ensure that we minimise this risk to
our patients when using any intravenous agent.
References
[ I ] THOMASDV. Propofol supports bacterial growth. British
Journal of Anaesthesia 1991; 66: 274.
[2] MILESAA, MISRASS. IRWIN JO. Estimation of bactericidal
power of blood. Journal qf Hygiene 1938; 38: 7 3 2 4 9 .
[3] Postsurgical infections associated with an extrinsically
contaminated intravenous anesthetic agent-California,
Illinois, Maine and Michigan, 1990. Morhidiry und Mortaliry
Weekly Report 1990; 39: 426-33.
[4] MCLEODGA, PACE N, INGLIS MD. Bacterial growth in
Propofol. British Journal of Anaesthesia 1991: 67: 665-6.