Ethylene Oxide Treatment Research Articles

Effect of the sterilization method on the performance of collagen type I on chronic wound parameters in vitro

In the treatment of chronic wounds, it is necessary to establish a physiological wound milieu to improve healing. Application of collagen as wound dressing has been described as beneficial as it possesses the ability to reduce elevated levels of proteases, cytokines, and free radicals. Consequently, a wide range of wound dressings based on collagen have been developed. Native collagen is susceptible to alterations because of influences during the production process; to minimize effects on the molecule itself collagen wound dressings are usually aseptically produced. Common sterilization methods (autoclaving, irradiation, and ethylene oxide (EtO) treatment) can induce changes in the protein chemistry and physical properties, potentially affecting the absorption rate, mechanical strength, or performance. In this study, we have evaluated the influence of gamma- and beta-irradiation as well as EtO sterilization on the binding capacity of collagen type I for selected proteases and cytokines associated with nonhealing wounds. Although a pronounced effect on the physical properties of the collagen was found, there was no significant loss in the binding affinity for polymorphonuclear elastase, matrix metalloproteinase-2, and interleukin-1beta, or in the antioxidant capacity.

Effects of Sterilizing Agents on the Biodegradation of a Bioplastic Material

A bioplastic material based on glucopolysaccharides and protein matrix sterilized by gamma radiation, ethylene oxide, moist and dry heat agents was studied. Biodegradability, measured by evaluation of carbon dioxide production and followed by electronic microscopy, was tested only in the irradiated and ethylene oxide-treated material because of the physical alterations experienced by the material submitted to moist and heat processes. The sterilized material showed excellent biodegradation in periods shorter than sixty days. Both sterilized and unsterilized materials showed excellent biodegradation. After gamma radiation and ethylene oxide treatment, the material maintained its mechanical properties of tensile strength and ultimate elongation without significant modifications. On the contrary, dry heat diminished elongation capacity by 80% and moist heat diminished tensile strength by 36%. Water absorption capacity of the material remained unchanged after all sterilization processes.

Bovine Bone Screws: Metrology and Effects of Chemical Processing and Ethylene Oxide Sterilization on Bone Surface and Mechanical Properties

We assess the effects of chemical processing, ethylene oxide sterilization, and threading on bone surface and mechanical properties of bovine undecalcified bone screws. In addition, we evaluate the possibility of manufacturing bone screws with predefined dimensions. Scanning electronic microscopic images show that chemical processing and ethylene oxide treatment causes collagen fiber amalgamation on the bone surface. Processed screws hold higher ultimate loads under bending and torsion than the in natura bone group, with no change in pull-out strength between groups. Threading significantly reduces deformation and bone strength under torsion. Metrological data demonstrate the possibility of manufacturing bone screws with standardized dimensions.

Comparison of the effects of sterilisation techniques on subsequent DNA profiling

It is important that contamination from extraneous DNA should be minimised on items used at crime scenes and when dealing with exhibits within the laboratory. Four sterilisation techniques (UV, gamma and beta radiation and ethylene oxide treatment) were examined for their potential to degrade contaminating DNA to such an extent that subsequent DNA profiling was impossible. This work indicated that the most successful technique to reduce DNA contamination was ethylene oxide treatment. Of the radiation techniques tested in this study, gamma was the most successful at eradicating DNA and UV radiation was the least. None of the contaminated samples treated with ethylene oxide and subsequently subjected to DNA analysis met the DNA profile criteria necessary for acceptance on the UK National DNA Database. Contaminated cotton swabs and micro-centrifuge tubes treated with ethylene oxide showed a marked decrease in amplifiable DNA post-treatment. Ethylene oxide treatment to sterile swabs and tubes did not significantly affect subsequent DNA analysis.

Development of a bacteriophage model system to investigate virus inactivation methods used in the treatment of bone allografts

Bone allografts are commonly used in a variety of surgical procedures, to reconstruct lost bone stock and to provide mechanical support during the healing process. Due to concerns regarding the possibility of disease transmission from donor to recipient, and of contamination of grafts during retrieval and processing procedures, it is common practice to sterilise bone allografts prior to issue for clinical use. It is vital that the sterilisation processes applied to allografts are validated to demonstrate that they achieve the required level of bioburden reduction, and by extension that validated models are used for these studies. Two common sterilisation protocols applied to bone allografts are gamma irradiation and ethylene oxide gas sterilisation, and there are currently no validated models available for measuring the anti-viral efficacy of ethylene oxide treatment with regard to bone allografts or readily useable models for assessing the anti-viral efficiency of gamma irradiation treatment. We have developed and validated models for both these sterilisation processes, using the bacteriophage varphix174, and utilised the models to measure the antiviral activity of the standard ethylene oxide and gamma irradiation sterilisation processes applied to bone allografts by the National Blood Service. For the irradiation model, we also utilised bacterial spores (Bacillus pumilus). Our results show that ethylene oxide sterilisation (which can only be applied to lyophilised grafts) inactivated > 6.1 log(10) of the model virus, and gamma irradiation (at 25 -40 kGy and applied to frozen allografts) inactivated 3.6 - 4.0 log(10) of the model virus and > 4 log(10) of the bacterial spores. Gamma irradiation at this dosage is therefore not in itself a sterilisation process with respect to viruses.

Characterization of methods for determining sterilization efficacy and reuse efficiency of oxygen biosensor multiwell plates

High-throughput screening (HTS) assays based upon fluorometric detection of oxygen consumption in microtiter plates were primarily developed for applications in drug discovery and ecotoxicology but have recently been adopted for use in microbial community-level physiological profiling assays (CLPP). The widespread use of oxygen biosensor systems for CLPP applications has, however, been hindered by the relatively high cost of oxygen biosensor reagent systems and limited access to microplate fluorometer instrumentation platforms. The ability to recycle and reuse oxygen biosensor system plates would expand their utilization for CLPP assays and other research applications in microbial ecology. Here, the efficacy and cost effectiveness of multiple procedures for sterilization of Oxygen Biosensor System™ (OBS; BD Biosciences) plates for reuse was evaluated. OBS plates were sterilized using ethylene oxide, ultraviolet radiation, and bleach treatments, then evaluated for biosensor response and plate life-cycle performance. Of the sterilization methods tested, ethylene oxide sterilization was most effective based on its low cost, high sterilization efficacy, and minimal impact upon OBS plate response.

Characterization of 75:25 Poly(l-lactide-co-ε-caprolactone) Thin Films for the Endoluminal Delivery of Adipose-Derived Stem Cells to Abdominal Aortic Aneurysms

Abdominal aortic aneurysms occur in 5-7% of men over the age of 60 and their incidence is rising. Current therapies remove the affected tissue or prevent blood flow through the aneurysm, but do not repair the underlying structural changes of the vascular wall. Adipose tissue derived stem cells (ADSCs) seeded on a biodegradable thin film and delivered endoluminally to the aneurysm site could potentially repair the vessel wall, preventing growth and rupture of the aneurysm. In this study, the mechanical and degradation properties of a novel 75:25 poly(l-lactide-co-epsilon-caprolactone) (PLCL) thin film, as well as, the effects of different surface structures on stem cell adherence and resistance to shear stress was investigated. It was possible to reproducibly create films of consistent physical properties. These films degraded approximately 50% in 6 month, which would be a sufficient time to allow cells to engraft in the aortic wall. Ethylene oxide treatment significantly increased the stiffness and yield stress of the films, which exhibit >700% elongation. Treatment of the films with NaOH and HCl induced the formation of surface texture on the films; however, this texture did not affect stem cell adherence or resistance to delamination by shear stress when compared to nontreated or fibronectin-coated films. These results indicate that PLCL thin films have a sufficient degradation time and mechanical strength to serve as a scaffold in vivo for ADSCs, and that ADSCs seeded on the thin film can withstand a range of physiologic shear stresses.

Sterilization of gentamicin containing collagen/PLGA microparticle composites

In order to achieve implants which provide sustained release of gentamicin, microparticles based on a 50/50 Resomer ® 503/Resomer ® 502H blend were combined with collagen in order to achieve their fixation and to utilize the favorable effect of collagen on wound healing. Ethylene oxide treatment as well as β- and γ-irradiation were tested for sterilization of the collagen/PLGA-microparticle composite. All methods resulted in a decrease of molecular weight and glass transition temperature of polymer raw material and microparticles. In addition, ethylene oxide treatment yielded aggregation of microparticles leading to a substantial increase in the initially liberated gentamicin dose. Furthermore, chemical changes of gentamicin after ethylene oxide sterilization could be identified using NMR spectroscopy. Despite a decrease in the molecular weight and glass transition temperature after irradiation, neither morphological changes of the composites nor changes regarding the gentamicin release profile from β- and γ-sterilized material were observed. Free radicals, which could only be detected in gentamicin drug substance and at marginal level in gentamicin-loaded MPs, disappeared within 4 weeks. Additional microbiological testing verified the microbiological activity of gentamicin liberated from β-sterilized composites. Storage of β-sterilized composite at 4 °C/35% r.h. for 3 months did not influence morphology, molecular weight, glass transition temperature, and release profiles of microparticles and composites. However, at 25 °C/60% r.h. and 40 °C/75% r.h. a marked decrease in molecular weight and glass transition temperature resulted. This effect was due to a higher humidity, water uptake into polymers, and subsequent hydrolysis of polymers and microparticles, which was more pronounced for RG 502H because of its hydrophilicity. Upon storage at 25 °C/60% r.h. and 40 °C/75% r.h. particles collapsed resulting in an increased gentamicin liberation. Thus, all sterilization techniques have their pros and cons, but based on drug release profile and chemical changes of gentamicin irradiation treatment appears to be more suitable for collagen/gentamicin-loaded PLGA microparticle composites.

Reconstruction of periprosthetic fractures of hip with cortical bone plates allografts

To observe clinical results for reconstruction of periprosthetic fractures of hip with cortical bone plates allografts by deep-freezing and ethylene oxide treatment. Seven patients with periprosthetic fractures of hip underwent cortical bone plates allografts by deep-freezing at -70 degrees C after being treatment of 48 degrees C ethylene oxide. And evaluate clinical outcome by examining T lymphocytes, Harris scores, X-rays photograph, and bone scintigraphy. There were not activity of immune rejection and infection in all patients. Harris scores of patients increased 21, 32, 40, 40 scores at 3, 6, 12, 24 months after surgery. T-lymphocytes, antibody and immunocomplex in blood was normal postoperation. X-ray film indicated that fracture was healed at 3 months and there was partially bone conjunction between allograft strut and host bone. There was incorporation of 85% allograft strut to host bone, and 15% allograft strut was partially absorbed at 12 months after surgery. The size of femur of host was added 3 mm to 5 mm, averaged 4.3 mm at 12 months postoperation. Density of 80% allograft plates was as same as host bone after remodeling and the absorbtion of 10% allograft plates stopped at 24 months after surgery. There was thick of nuclein in the area of allograft cortical bone plates by bone scintigraphy examination at 3 months postoperation, and the thick of nuclein was stronger at 6, and 12 months after surgery. Allograft cortical bone plates by deep frozen at -70 degrees C after being treatment of 48 degrees C ethylene oxide is suitable for mechanical fixation and biological bone transplantation, and it can increase bone reservation, augment strength of femur once the allograft strut incorporates to host bone, and avoid removing metal implant in second operation when being applied into reconstruction femoral fracture in joint replacement.

Vibrational spectroscopy of ultra-high molecular weight polyethylene hip prostheses: influence of the sterilisation method on crystallinity and surface oxidation

Due to its high strength and low creep, ultra-high molecular weight polyethylene (UHMWPE) has been used for 30 years in the replacement of damaged articulating cartilage for total joint replacement surgery. In this study, micro-Raman spectroscopy was used to investigate the effects of the sterilisation method (gamma and ethylene oxide (EtO) treatment) on the crystallinity changes of UHMWPE acetabular cups. The crystallinity of the cups was evaluated by micro-Raman spectroscopy coupled to the partial least square (PLS) regression as a function of the inner surface position. Unworn gamma-sterilised cups were found to be significantly more crystalline than the EtO-sterilised ones. No significant differences were observed between the crystallinity values of worn (in in vitro tests) and unworn cups for each type of sterilisation, showing that the changes in surface crystallinity were mainly caused by irradiation rather than by mechanical friction during the in vitro wear tests. These results were discussed in relation to gravimetric measurements, which revealed at the end of the in vitro tests, a higher mean weight loss for the EtO-sterilised cups than for the gamma-sterilised ones. No significant amounts of oxidative degradation products were detected by IR spectroscopy in the inner surface of the EtO-sterilised worn and unworn cups. Regarding the gamma-sterilised cups, the oxidation level appeared to be slightly higher in the centre of the worn cups than on their borders.

DNA damage induced by gamma-radiation in combination with ethylene oxide or propylene oxide in human fibroblasts

To estimate the effects of interaction of gamma-rays and an epoxide, cell survival and induction of DNA double-strand breaks (DSBs) following combined exposure to ionizing radiation and ethylene oxide (EtO) or propylene oxide (PO) were studied in human fibroblasts. Two treatment protocols were applied: (a) the cells were pre-exposed to different doses of gamma-rays and then treated with epoxide, and (b) the cells were pretreated with epoxide and then exposed to different doses of gamma-rays. Here we show that order of the treatment did not play a role in cell survival and that the effect of combined exposure on cell killing was additive for both epoxides. As to DNA DSBs induction, however, a difference dependent upon the order of the treatment was observed. While EtO or PO treatment followed by gamma-rays exposure led to an increased number of DSBs at higher gamma-ray doses (2–3 Gy), no significant increase of DSBs was detected after the opposite order of the treatment (gamma-ray exposure followed by EtO or PO treatment).

Effects of the sterilisation method on the wear of UHMWPE acetabular cups tested in a hip joint simulator

Ultra-high molecular-weight-polyethylene is the most commonly used bearing material in total joint replacement. Wear of polyethylene is a serious clinical problem that limits the longevity of orthopaedic implants. Information on degradative changes in the material properties and on the methods used for the sterilisation of polyethylene may help in the selection process of orthopaedic implants with the best wear resistance. This study was performed to investigate the effects of the sterilisation method (gamma irradiation and ethylene oxide treatment) on the wear and on the changes in physical properties of polyethylene acetabular cups. At this purpose, gamma-sterilised and ethylene oxide (EtO)-sterilised acetabular cups were tested against CoCr femoral heads in a hip joint simulator run for 5 million cycles in bovine calf serum. The crystallinity of the cups was evaluated by micro-Raman spectroscopy as a function of the inner surface position. The partial least square calibration was used to correlate the Raman spectra with the crystallinity of the polymer measured by differential scanning calorimetry. The analysis performed on soak control acetabular cups demonstrated that the gamma-sterilised cups are significantly more crystalline than the EtO-sterilised ones. The mean crystallinity values obtained for the gamma-sterilised and EtO-sterilised soak control cups were 65.0% and 63.4%, respectively. Weight loss measurements revealed that the gamma-sterilised acetabular cups exhibited a lower wear rate than that by EtO-sterilised. The Raman results obtained on gamma-sterilised and EtO-sterilised acetabular cups showed that the changes in surface crystallinity were mainly caused by irradiation rather than by the mechanical friction during the in vitro tests.

The effect of solvent extraction and sterilization procedure on the tissue response to Dacron ® velour

The effects of solvent extraction and sterilization procedure on tissue response to Dacron ® velour were studied in a canine model, using histomorphometrical techniques. The solvents used for extraction of low molecular weight moieties were either ethanol or water; the sterilization techniques examined were ethylene oxide (ETO) treatment, steam sterilization, and radiofrequency glow discharge (RFGD) treatment. The effect of the sterilization procedure was most marked in the outermost regions (velour) of the implant; no sterilization effects were determined in the capsule or the knitted regions. Velour in the steam sterilized implants had the smallest blood vessel dimensions compared with those that were treated with ETO or RFGD. The effects of different extraction methodologies appeared to be more significant than sterilization effects and were detected further into the implant. That is, not only were extraction effects detected in the capsule and velour, they were also detected in the outer knitted region. Extraction with water resulted in histological responses considered more biologically desirable (thinner capsule, lower giant cell presence, and larger blood vessel diameters) than responses to extraction with ethanol. Neither extraction nor sterilization effects were detected in the inner layers of the knitted region, which were adjacent to the adhesive.

Failure of ethylene oxide to sterilize extracted human teeth

Objectives. The purpose of this study was to determine if ethylene oxide could sterilize extracted human teeth to be used in research. Methods. An occlusal preparation was cut in freshly extracted molars, and a small hole was drilled into the pulp chamber. A suspension of Bacillus subtilis (globigii) endospores, the standard biological monitor used for ethylene oxide sterilization, was injected into the pulp cavity, and the pulp cavity access was filled with composite material and sealed with a light-cured sealant. The teeth were exposed to either a 30°C or 63°C ethylene oxide sterilization process. Following exposure, the teeth were aseptically split and cultured to reveal viable spores. Results. Sixty-four percent of the teeth exposed to “cold” ethylene oxide treatment and 80% of the teeth exposed to the “warm” treatment still contained viable spores. Significance. Ethylene oxide cannot be relied on to sterilize extracted human teeth. Therefore, before they are used in research, other methods should be used to ensure killing of bloodborne pathogens that may be present within the teeth.

Evaluation of a series of tyrosine-derived polycarbonates as degradable biomaterials.

A series of four polycarbonates derived from the ethyl, butyl, hexyl, and octyl esters of desaminotyrosyl-tyrosine was prepared by condensation polymerization. The resulting polymers had weight average molecular weights ranging from 120,000-450,000, and their chemical structure was confirmed by elemental analysis, nuclear magnetic resonance, and Fourier transform infrared spectroscopy. The polycarbonates were evaluated as degradable biomaterials. Their surface properties were determined by electron spectroscopy for chemical analysis, attenuated total reflectance-Fourier transformed infrared spectroscopy, and contact angle measurement. The degree of surface hydrophobicity was related to the length of the alkyl ester pendent chain. The tensile properties were dependent on the chemical structure of the polymers: For thin, solvent cast film specimens, the tensile modulus varied from 1.2-1.6 GPa, and the strength at break from 60-220 MPa. The degradation of polymeric films was followed in vitro by measuring changes in mechanical strength for up to 40 weeks, and the decrease in molecular weight and changes in surface chemistry for up to 80 weeks. The length of the pendent chain affected the degradation behavior and strength retention; the polymers with short pendent chains were more readily hydrolyzable. For sterilization, ethylene oxide treatment was less destructive, as judged by molecular weight retention, than gamma-irradiation. Spin-cast films of all tested polycarbonates were not cytotoxic toward cultured rat lung fibroblasts. The cell response was influenced by the chemical structure of the polymer. The least hydrophobic polycarbonate (having a short ethyl ester pendent chain) was a more stimulating substrate for cell growth than the more hydrophobic polymers (carrying longer alkyl ester pendent chains).

Evaluation of polyphosphates and polyphosphonates as degradable biomaterials

A series of polymers, bisphenol A-based poly(phosphoesters), were evaluated as degradable biomaterials. Degradation was observed for the four polymers studied under both in vitro and in vivo conditions. The rate of degradation was affected by polymer side-chain structure and correlated with the swelling behavior. The ethyl side-chain polymers absorbed more water than their phenyl counterparts. Among the sterilization methods, UV irradiation followed by antibiotic treatment was the most suitable, as steam autoclave and ethylene oxide treatments altered the properties of several of the poly (phosphoesters). Tissue response to the poly(phosphoesters) in rabbits was characterized by minor encapsulation and slight or no lymphocyte, giant cell, or macrophage activity. No evidence of edema or necrosis was found. The elastic moduli of these materials varied from 488 MPa for poly(bisphenol A-ethylphosphate) (BPA/EOP) to 627 MPa for the more rigid poly(bisphenol A-phenylphosphonate) (BPA/PP). The ultimate strength, modulus, and energy to failure of BPA/PP were lower than those of similarly compression molded high-molecular-weight poly(L-lactic acid) (PLLA).

Induction of glutathione- S-transferase and heat-shock proteins in rat liver after ethylene oxide exposure

Defense mechanisms in rat liver against depletion of glutathione (GSH) and cellular injuries induced by ethylene oxide (EO) were studied. Rats were exposed to EO under either high dose (1300 ppm for 4hr, once) or low dose (500 ppm for 6hr, three times a week for 6 weeks) conditions. The hepatic content of GSH decreased dramatically after EO treatment, probably due to detoxication of EO. After the high dose treatment the hepatic GSH content fell by 90% of the control values but recovered within 10 to 15 hr. EO reacts directly with a variety of cellular macromolecules but all rats survived the exposure. Since the metabolites of EO are ethylene glycol and GSH-conjugates, the enzymatic activities of epoxide hydrolase and glutathione- S-transferase (GST) were determined. Only GST activity was found to occur after low dose chronic exposure. The defense mechanism at mRNA level was investigated using probes for GST and several heat-shock proteins (hsps). Enhanced accumulation of GST mRNA was detectable during the recovery period of rats after both high and low dose exposure to EO. Interestingly, both hsp32 (< 40-fold) and hsp90 (< 3-fold) mRNA increased after high dose exposure but the mRNA level of one of the major heat-shock proteins, hsp70, did not change under these conditions. Diethylmaleate, which is known to be a GSH depleter in liver, induced hsp32 mRNA only in rat liver, while hsp70 and hsp90 mRNA levels did not change when GSH was depleted. These results suggest that individual heat-shock proteins are induced in different ways under unphysiological conditions such as EO exposure.

Biological qualities of saliva sterilized by filtration or ethylene oxide treatment

Stimulated whole saliva was centrifuged and sterilized either by filtration in the presence or absence of dithiothreitol (DTT) or by ethylene oxide treatment. Filtration was accomplished by 2 different techniques: conventional positive pressure filtration through prefilters and membranes with decreasing pore sizes and tangential flow microporous filtration. The sterile saliva samples were analyzed for enzyme activities of phosphatases, esterases, glucuronidase, and lysozyme. The samples were also tested by polyacrylamide electrophoresis, isoelectric focusing and for IgA concentration. Moreover, the samples were tested for support of bacterial growth of strains from the genera Actinomyces, Lactobacillus, Neisseria, Rothia and Streptococcus after the addition of glucose at a final concentration of 0.5%. The samples treated with ethylene oxide appeared to be more affected by this treatment than samples treated otherwise, as demonstrated by the electrophoretic analyses and analyses of enzyme activities. Ethylene oxide-treated saliva also was the least favorable saliva sample to support growth of bacteria. The positive pressure technique gave more rapid filtration than the tangential flow technique. DTT treatment of saliva facilitated filtration and, in contrast to ethylene oxide treatment, it affected the qualities of saliva only to a minor extent compared with the original saliva sample.

Sterilization Mode Effect on the Hydration Rate of Calcium Sulfate Hemihydrate

ABSTRACTPlaster bandages have been found to support bacteria. The sterilization of plaster (calcium sulfate hemihydrate) for use as an implant biomaterial is, therefore, a primary concern. Heat sterilization can significantly alter material properties and the kinetics of plaster hydration. A study has been done of the effects of sterilization by gamma radiation and by ethylene oxide on the hydration rate of plaster. Three hemihydrate bases with differing water requirements and hydration rates were sterilized by each method. Adiabatic temperature rise was used to follow hydration. Five outcomes were measured and analyzed. These outcomes include the induction period, maximum rate of temperature rise (MRR), time to MRR, total temperature rise, and time to maximum temperature. Neither sterilization mode was found to significantly alter the hydration process of any of the plaster bases. From a material property standpoint, either gamma radiation or ethylene oxide treatment is preferred to heat sterization of calcium sulfate hemihydrate.

Comparative study of micronucleus assay and chromosomal aberration analysis in V79 cells exposed to ethylene oxide

Micronucleus formation and chromosomal aberration (CA) in V79 cells were compared for their sensitivity in response to ethylene oxide (EtO) treatment. The results indicate that EtO exposure for 30 min induced CAs in V79 cells at the concentration of 3,500 ppm or higher. A statistically significant difference (P less than 0.01) was found between treated and control groups at all concentrations tested based on percent aberrant cells. The increase was dose-dependent with a correlation coefficient of 0.88. The aberrations found include chromatid and isochromatid breaks, fragments, minutes, and exchanges. Results of the micronucleus assay both in mononucleated and binucleated cells showed a slight but not statistically significant increase in micronuclei with doses between 457 to 4115 ppm. At the highest concentration tested (12344 ppm) EtO caused a significant increase in the micronucleus frequency (P less than 0.05).