Eo Residue in Medical Devices 03
Eo Residue in Medical Devices 03
Eo Residue in Medical Devices 03
net/publication/10660163
CITATIONS READS
50 7,666
7 authors, including:
Anne D Lucas
U.S. Department of Health and Human Services
71 PUBLICATIONS 1,020 CITATIONS
SEE PROFILE
All content following this page was uploaded by Anne D Lucas on 11 December 2017.
Abstract: Ethylene oxide (EO) gas is commonly used to sterilize medical devices. The
amount of residual EO remaining in a device depends partly on the type and size of polymeric
material. A major concern is the amount of residue that may be available in the body. With
the use of the method described by AAMI for headspace analysis of EO residues, different
polymers and medical devices subjected to different numbers of sterilization cycles were
examined. Next, the effect of various extraction conditions and extraction solutions on these
polymers and medical devices was evaluated. The results showed different polymers desorb
EO differently. One polyurethane (PU 75D) had much higher EO residue than a different
polyurethane (PU 80A). Repeated extraction of the PU 75D was necessary to quantify total EO
residue levels. Different extraction solutions influence the amount and reproducibility of EO
detected, whereas multiple resterilizations showed no difference in amount of residual EO.
Bioavailability of EO was estimated by extracting the devices and polymers in water. Com-
parison of total EO residues to EO that was bioavailable showed no difference for some
polymers and devices, while others had an almost eightfold difference. Some standard bio-
compatibility tests were run on extracts and devices, but no significant effects were observed.
© 2003 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater 66B: 548 –552, 2003
device containing EO in an aqueous solution at 37 °C for heating to 100 °C for 60 min, followed by GC residue
24 h. Following the analytical work, some biocompatibility analysis of the headspace gas. Samples were repeatedly
tests were conducted on EO sterilized polymers and the heated and analyzed, with nitrogen purge or evacuation be-
aqueous extracts of EO sterilized devices. tween cycles, until EO levels approached the limit of reliable
quantitation (approximately 5 g/g).
To evaluate bioavailability of EO residues, water, culture
MATERIALS AND METHODS media [RPMI-1640 with L-glutamine and 10% fetal bovine
serum (FBS)], and cottonseed oil were compared as extrac-
Materials tion solutions. EO was added to each solution (500 g) and
Two different polyurethanes, Pellethane 2363-75D (PU 75D) kept at 37 °C for 24 h.9 The following day, 1-ml aliquots were
and 2383-80A (PU 80A), and Nylon 66 were tested. Material removed and analyzed. Based on the results of the three
sheets 0.5 mm thick were cut into 10 ⫻ 45-mm strips with extraction solutions, water was deemed best, and all of the
five strips per specimen sample. Two types of electrophysi- materials were subsequently extracted in 13 ml of water for
ology diagnostic electrode catheters were also tested. Both 24 h at 37 °C with 1 ml removed for GC residue analysis. The
types were from the same manufacturer (Bard), made of hollow and solid catheter pieces used for liquid extraction had
cross-linked polyurethane (personal communication), and ap- a final surface area to extraction solution volume ratio of 1.96
peared to be identical except for the connectors. However, cm2/ml. PU 75D, PU 80A, and Nylon 66 had a 3.5 cm2/ml
one catheter had embedded wires (designated solid or S) in surface area to volume ratio.8
the shaft, whereas the other one was hollow with removable
Biocompatibility Testing
wires (hollow or H). The wires in the hollow catheter samples
were removed before sterilization. The catheters were cut into Apoptosis and Cytotoxicity. In one series of tests, 1 ml
45-mm-long sections with nine sections composing a speci- of the water extracts for each material was placed into cell
men sample. culture with 2-ml Jurkat cells (a human lymphoma cell,
ATCC CRL 8163) to test viability and appearance of apo-
EO Sterilization ptotic cells. In addition, EO standards (final concentration in
Materials were sterilized on the weekend at the NIH Clinical cell culture 0.2 to 83 g/ml) were prepared in water and
Center with the use of 10% EO at 54 °C, 65% RH for 130 tested with these cells. Twenty-four hours after adding the
min, followed by 12 h aeration at 132 °C. On Monday, after sample or EO standard, an aliquot of the cells was removed
2 days at room temperature, the samples were retrieved and and analyzed using a flow cytometer and LYSIS I-I software
stored at ⫺ 70 °C until prepared for analysis. For resteriliza- (FACScan, Becton Dickenson, San Jose, CA), as previously
tion studies, samples were left at room temperature for 5 days described.10,11 Cells were analyzed according to the side-
before resterilization with EO. Specimens were EO sterilized scattering profile (proportional to cellular granularity) verses
0, 1, or 5 times. the forward-scattering profile (proportional to the cellular
cross sectional area). Changes in the cell populations were
Analytical Method evaluated by gating on the normal cell population and com-
paring to the test groups over time. Apoptotic cells exhibit a
The EO stock standard used in this study was 50 mg/ml in decrease in forward scatter (reduced cell size) and an increase
methanol (Suppelco 4-8838). The standard was then diluted in side scatter (increased granularity) and can readily be
in water or methanol as needed. EO levels were determined quantified with the use of a flow cytometer.10 –13
using the ISO method7. Headspace sampling is a method of
introducing volatile components, such as EO, from a liquid or Cytotoxicity Testing. The polymer PU 75D, which had
solid sample into a gas chromatograph. The vial is heated, the highest EO residue, was tested for its effect on fibroblasts
allowing the volatile compounds to go into the air (or head- in culture. EO levels for this particular lot of PU 75 D were
space) above the liquid or solid sample. After heating, a 1540.7 g/g EO for exhaustively extracted and 458 g/g in
gastight syringe is used to remove a portion of the air from the water extracts. The protocol was conducted according to
the headspace and inject the sample directly into the gas ANSI/AAMI/ISO14 using extracts and according to ASTM F
chromatograph. A Hewlett-Packard headspace auto sampler 81315 using direct contact tests. The PU 75D samples were
(HP 7694 oven 100 °C, loop 105 °C, vial equilibration time either used immediately after retrieval following the EO
15– 60 min, pressure: 0.5 min, loop fill: 0.15 min, loop sterilization and aeration protocol (detailed in EO sterilization
equilibration time 0.05 min), a HP gas chromatograph 5890 section) or were kept frozen at ⫺70 °C until use. The control
series II (inlet 105 °C, 30 m ⫻ 0.32 mm Omega-wax 320 PU 75D needed to be sterilized by a means other than EO for
capillary column; 30 °C 5 min, 20 °C/min to 100 °C, hold 15 comparison; so control samples were sterilized by autoclav-
min) and an FID detector (220 °C) were used. ing. Samples were cut to size before the sterilization cycles.
L929 fibroblast cells were grown to a confluent monolayer in
Extraction Method
100 ⫻ 15-mm culture dishes with RPMI 1640 medium con-
Samples were analyzed for total EO concentration. Solid taining 10% FBS. The medium was removed prior to addition
specimens were sealed in a vial and thermally extracted by of the sample or extracts.
550 LUCAS ET AL.
Exhaustive Extraction
RESULTS AND DISCUSSION Figure 2. Cumulative EO thermally extracted from PU 75D, Nylon 66,
and solid catheter pieces. Samples were thermally extracted eight
Resterilization times at 60 min at 100 °C then analyzed until EO levels were at the
limit of detection. Both PU 80A and the hollow catheter had very low
The amount of EO varied greatly for the different samples. levels of EO (less than 10 g/g and 10.4 g/g, respectively; data not
However, the number of sterilization cycles had little influ- shown).
RESIDUAL ETHYLENE OXIDE IN MEDICAL DEVICES 551