|Year : 1993 | Volume
| Issue : 1 | Page : 20-22
The efficacy of acetone in the sterilisation of ophthalmic instruments
Vinay Agrawal, Savitri Sharma
L.V. Prasad Eye Institute, Hyderabad, India
L.V. Prasad Eye Institute, Road No. 2, Banjara Hills, Hyderabad - 500 034
Source of Support: None, Conflict of Interest: None
Acetone has been considered a quick, effective and less expensive chemical sterilising agent and continues to be used by ophthalmic surgeons, at least in developing countries. Its utility however has been questioned recently. This study was designed to assess the efficacy of acetone against Pseudomonas aeruginosa, Bacillus subtilis and Aspergillus flavus present on ophthalmic surgical instruments (forceps, sutures). The instruments were contaminated by immersion in standard suspensions of the organisms and thereafter were either unwashed (group-I), washed and dried (group-II) or only washed (group-III) before immersion in acetone. The exposure to acetone was kept at 3, 10 and 20 minutes in each group. The results showed that acetone could eliminate Pseudomonas (vegetative bacteria) after 10 minutes exposure in unwashed group and 3 minutes exposure in washed groups. It was ineffective against spore bearing bacteria (B. subtilis) and fungus (Aspergillus flavus) even after 20 minutes of exposure
Keywords: Acetone sterilisation; ophthalmic instruments
|How to cite this article:|
Agrawal V, Sharma S. The efficacy of acetone in the sterilisation of ophthalmic instruments. Indian J Ophthalmol 1993;41:20-2
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Agrawal V, Sharma S. The efficacy of acetone in the sterilisation of ophthalmic instruments. Indian J Ophthalmol [serial online] 1993 [cited 2020 Apr 5];41:20-2. Available from: http://www.ijo.in/text.asp?1993/41/1/20/25630
Sterilisation of surgical instruments employing chemicals is a popular method. The conventional methods of sterilisation such as autoclaving may blunt sharp instruments. Ethylene oxide sterilisation has emerged as a highly effective alternative to autoclaving for articles that may be damaged by heat. However, the equipment is expensive and the sterilised articles require aeration for several days. Similarly, dry heat sterilisation in a hot air oven is also time consuming. This has led to the suggestions for use of chemical methods for sterilisation. Glutaraldehyde (2%) and cetrimide (0.5 %) are some of the chemicals that are available for such use. For these to be effective several hours of contact is required followed by careful washing of the instrument, prior to usage . Acetone seemed to overcome these drawbacks and was considered a quick , effective and less expensive method of chemical sterilization ,,. It may also be useful as an alternative to sterilise instruments accidentally contaminated during surgery or required at short notice.
The efficacy of acetone claimed from previous experience has been in question. Since discrepancy in the design of the experiments conducted by the various authors may be a possible factor, we planned to evaluate the efficacy of acetone as a sterilising agent for ophthalmic instruments.
| Materials and methods|| |
Sterile toothed forceps and 10-0 Nylon sutures with curved cutting needles were used as test articles. Two bacterial isolates of Pseudomonas aeruginosa, B. subtilis and one fungal isolate, Aspergillus flavus were used as challenge strains. All strains were isolated from corneal ulcer patients attending the out patient clinic. Pure (99.5%) commercial grade Acetone (Merck) was used as the test reagent.
Pseudomonas aeruginosa was grown overnight in Tryptone Soya Broth (TSB) and a0.1% inoculum was made in fresh TSB. This was grown at 35°C without agitation to a concentration adjusted to 1.4x 10 5 cfu/ml [optical density of 0.06 at 420nm on a spectrophotometer, ELICO India]. The final concentration was verified in serial dilutions by spread plate method .
Bacillus subtilis strain was grown on nutrient agar for 3 days allowing for sporulation to occur which was verified by Gram's stain. Standardised suspensions of 1.6 x 10 5cfu/ml was made by suspending the culture in 0.85% saline, adjusting the optical density to 0.03 and verifying the concentration by serial dilutions.
Aspergillus flavus was grown on Sabouraud's dextrose agar for 48 hours at 25-27°C. The surface spores from the growth were harvested into 0.85% saline and a concentration of 2 x 10 5 cfu/ ml (OD-0.02) was achieved as described forBacillus subtilis.
The study design used for the performance of the experiments is described in [Figure - 1]. The test instruments and sutures were immersed in the microbial suspension for 10 seconds and divided into three groups. In group I the test articles were immersed in the bacterial/fungal suspension for 10 seconds and then immersed directly into acetone. For group II after removal from the microbial suspension the instruments were washed with sterile distilled water and wiped with sterile towel before immersion in acetone. For group III only washing (without drying) was carried out. The test articles in each group were immersed in 500 ml of acetone kept in sterile beaker for 3,10 or 20 minutes. They were then removed, allowed to air dry (approximately 10 seconds) and placed in culture tubes. The culture media used were Brain Heart infusion broth and Sabouraud's dextrose broth for articles treated with bacteria and fungus respectively. The acetone used for the experiment was unchanged through one set of experiments to simulate its likely mode of use in clinical situation. The controls-were similarly treated using distilled water instead of acetone. The test instruments or sutures placed in the media were incubated at 37°C and 27°C for bacteria and fungus respectively. The media were examined for turbidity (growth) at the end of 24 and 48 hours. A subculture on nutrient agar for bacteria and Sabouraud's dextrose agar for fungus was performed whenever the turbidity of liquid medium was not obvious or contamination was suspected 
| Results|| |
The viability of all the three test organisms used for the experiments after exposure to acetone is depicted in [Figure - 2]. Pseudomonas aeruginosa was not eliminated by 3 minute exposure to acetone in group I i.e. when the articles were immersed in acetone directly after removal from the microbial suspension without any washing or wiping. However, the organisms were eliminated after exposure to acetone for 10 and 20 minutes in the same group. After washing and wiping (Group II) and only washing (Group III) following removal from microbial suspension and prior to immersion in acetone, a 3 minute exposure to, acetone was adequate, as were 10 and 20 minutes in eliminating the bacteria.
None of the articles contaminated with Bacillus subtilis and Aspergillus flavus were rendered sterile even after 20 minute exposure to acetone in any of the 3 groups. In all the groups there was significant turbidity of the broth and pellicle formation after 24 hours itself Subculture of the turbid broth after 48 hours of incubation confirmed the presence of the test organisms.
All the controls were positive only for the organisms tested.
| Discussion|| |
Chemical sterilisation of surgical instruments with acetone has been reported to have varying efficacy . Though in use since 1956 1 no scientific data is available on its efficacy.
A study by Sudhakar et al 2- concluded that acetone is the `best available instant sterilisation agent'. However, they have used solid media to test for the viability of organisms on the ophthalmic instruments. The method used will obviously not allow the growth of all organisms present on the instrument. The study by Madhavan et a[ 4], was the first to question the efficacy of acetone. They concluded that acetone was not safe for use as a chemical sterilisation agent. They used only the 3 minute exposure to acetone for vegetative bacteria in their experiments and did not test acetone against fungi.
Our results [Figure - 2] indicate that acetone is effective against vegetative bacteria when the articles were washed and dried before immersion in acetone and kept for 10 minutes. However, acetone was ineffective against spore bearing bacteria as well as fungi even after 20 minutes of exposure. The washing and drying steps did not affect the outcome.
It is not possible to comment from this study regarding the number of organisms that were actually present on the articles when they were placed in acetone. However, the number of organisms was likely to be extremely low atleast in group II and III (owing to washing) and probably not more than what may normally be present on a contaminated instrument.
We are aware of surgeons who have been using acetone as sterilising agent for years "without any problem". Our contention is that in the event of post operative infection the source may not be traced to acetone. Post operative infections can be from multiple sources and it requires an extremely well planned investigation to trace the source. More often, it remains undetected.
Sterilisation is defined as "eradication of both vegetative and spore forms of microorganisms". Inability to kill spores cannot allow an agent to be called a sterilising agent, more so for use with critical items. Surgical instruments which are introduced beneath the surface ofthe body (forceps, scissors, blades, needles etc) are regarded as critical items. They need to be free of microbial spores in addition to vegetative bacteria, fungi and viruses as defined by Spaulding'.
The use of acetone is thus an unacceptable compromise in a clinical situation where the actual load of organisms and the type of organisms cannot be predetermined. It is amply clear that acetone is not a safe chemical sterilisation agent and its use as one in any type of situation cannot be a recommended practice.
| Acknowledgement|| |
This work was supported by a grant from the Hyderabad Eye Research Foundation, Hyderabad, India
The authors acknowledge the technical assistance of Mr. D. Venkateshwara Rao and Mr. K. Sekhar Reddy and thesecretarial assistance of Ms. K. Malini.
| References|| |
Drews RC. Acetone sterilization in ophthalmic surgery. Ann Ophthalmol. 9: 781-784, 1977.
Sudhakar A and Kakadia J. Clinical and microbiological assesment of acetone as a chemical sterilization agent in microsurgery. Proc. 40th All India Ophth Soc. 322-325, 1990.
Fechner PU and Alpar JJ. Microscopes, instruments, acetone, disinfection, Sheet's glide. In, Intraocular lenses, Jaypee Bros; New Delhi, 213, 1988.
Madhavan HN, Rao BS and Vardarajan J. Acetone sterilisation of ophthalmic instruments. Nethralaya insight, 9 & 10: 7 - 9, 1992.
Brown R, Proxton IR and Wilkinson JF. Centrifuges, Colorimentars and bacterial counts. In practical microbiology: Colle JG, Duguid JP, Fraser AG, Marmion BP (eds). Churchill Livingstone, London. 240-247, 1989.
Colle JG and Milles RS. Tests for identification of bacteria. In, Practical medical micrbiology: Colle JG, Duguid JP, Fraser AG; Marmion BP (eds) , Churchill Livingstone, London. 141-160, 1989.
Spaulding EH, Lundy KR and Turner FJ. Chemical disinfection of medical and surgical materials. In Block SS (ed): Disinfection, sterilization and preservation; Lea and Febiger, Philadelphia, 1977.
[Figure - 1], [Figure - 2]
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