The antimicrobial effect of acetic acid—An alternative to common local antiseptics?

The antimicrobial effect of acetic acid—An alternative to common local antiseptics?

burns 35 (2009) 695–700 available at www.sciencedirect.com journal homepage: www.elsevier.com/locate/burns The antimicrobial effect of acetic acid—...

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burns 35 (2009) 695–700

available at www.sciencedirect.com

journal homepage: www.elsevier.com/locate/burns

The antimicrobial effect of acetic acid—An alternative to common local antiseptics? H. Ryssel a,b,*, O. Kloeters a,b, G. Germann a,b, Th. Scha¨fer a,b, G. Wiedemann a,b, M. Oehlbauer a,b a b

Department of Hand, Plastic and Reconstructive Surgery and Burn Center, BG Trauma Center Ludwigshafen, Ludwigshafen, Germany Department of Plastic and Hand Surgery – The University of Heidelberg, Heidelberg, Germany

article info

abstract

Article history:

Acetic acid has been commonly used in medicine for more than 6000 years for the disin-

Accepted 17 November 2008

fection of wounds and especially as an antiseptic agent in the treatment and prophylaxis of the plague. The main goal of this study was to prove the suitability of acetic acid, in low

Keywords:

concentration of 3%, as a local antiseptic agent, especially for use in salvage procedures in

Acetic acid

problematic infections caused by organisms such as Proteus vulgaris, Acinetobacter baumannii

Antimicrobial power

or Pseudomonas aeruginosa.

Bacteria

This study was designed to compare the in vitro antimicrobial effect of acetic acid with

Burns

those of common local antiseptics such as povidone–iodine 11% (Betaisodona1), polyhex-

Topical antiseptics

anide 0.04% (Lavasept1), mafenide 5% and chlohexidine gluconate 1.5% cetrimide 15% (Hibicet1). Former studies suggest the bactericidal effect of acetic acid, but these data are very heterogeneous; therefore, a standardised in vitro study was conducted. To cover the typical bacterial spectrum of a burn unit, the following Gram-negative and Gram-positive bacterial strains were tested: Escherichia coli, P. vulgaris, P. aeruginosa, A. baumannii, Enterococcus faecalis, Staphylococcus epidermidis, methicillin-resistant Staphylococcus aureus (MRSA) and b-haemolytic Streptococcus group A and B. The tests showed excellent bactericidal effect of acetic acid, particularly with problematic Gram-negative bacteria such as P. vulgaris, P. aeruginosa and A. baumannii. The microbiological spectrum of acetic acid is wide, even when tested at a low concentration of 3%. In comparison to our currently used antiseptic solutions, it showed similar – in some bacteria, even better – bactericidal properties. An evaluation of the clinical value of topical application of acetic acid is currently underway. It can be concluded that acetic acid in a concentration of 3% has excellent bactericidal effect and, therefore, seems to be suitable as a local antiseptic agent, but further clinical studies are necessary. # 2008 Elsevier Ltd and ISBI. All rights reserved.

Local infection and burn-wound sepsis are still some of the most severe problems in the treatment of thermally injured patients. Morbidity and mortality of burn patients is highly correlated to the incidence of wound infection and its sequelae [1].

Colonisation is the first step to infection, which can not only cause burn-wound sepsis but also induces immune responses, that leads to cytokine release, the entire cascade of acute-phase reaction and subsequent multi-organ failure.

* Corresponding author at: Department of Hand, Plastic and Reconstructive Surgery and Burn Center, BG Trauma Center Ludwigshafen, Ludwig-Guttmann-Str. 13, D-67071, Ludwigshafen, Germany. Tel.: +49 621 6810 2942; fax: +49 621 6810 2609. E-mail address: [email protected] (H. Ryssel). 0305-4179/$36.00 # 2008 Elsevier Ltd and ISBI. All rights reserved. doi:10.1016/j.burns.2008.11.009

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burns 35 (2009) 695–700

At present, many different agents, such as silver nitrate, mafenid acetate or povidone–iodine, are applied to burn wounds in an attempt to reduce bacterial colonisation. However, a controversial discussion concerning the selection of the ‘ideal’ antiseptic treatment is still ongoing. At a time when bacterial resistance to antibiotics is a matter of increasing concern, the value of ‘household remedies’, such as acetic acid, as salvage procedures needs to be re-evaluated and discussed [2]. Therefore, the bactericidal effect of acetic acid against common bacterial strains found in our burn patient unit was examined to evaluate the suitability of low concentration acetic acid as a local antiseptic agent. The use of acetic acid in the treatment of wound infection dates back to 1916 when Taylor found that application of a 1% solution to war wounds led to the elimination of Bacillus pyocyaneus after 2 weeks [3]. Because of the different patterns of injury, the data in this report are very heterogeneous and the grade of bacterial contamination of the different wounds is also unknown. In 1968, Phillips reported about the use of a 5% solution of acetic acid in superficial wounds [4]. Even in this study the data are very heterogeneous because patients with ulcers, burns, Kaposi’s sarcoma, skin lacerations, etc., are included in the study group. Another problem in this study is the parallel use of antibiotics and other local antiseptics such as chlorhexidine [4]. Phillips focussed on P. aeruginosa, but the germs in a burn unit are quite different and the bacterial load is also unknown. With his solution of 5% acetic acid, he reported side effects of severe itching and pain [4]. Sloss et al. [5] reported about eight patients and acetic acid concentrations between 1% and 4%. The number of patients studied was small and the population heterogeneous. He also reported on itching and pain as side effects in the use of higher concentrations of acetic acid [5]. The data from the earlier studies suggest an antimicrobial effect of acetic acid [3–5]. However, all these studies involve small number of cases and demonstrate very heterogeneous data; therefore, further trials like the present study are necessary to re-evaluate the effect of acetic acid. The effective concentration seems to be between 1% and 5% [3–5], with the problem that higher concentrations are associated with severe pain and itching [4,5]. Therefore, we used a concentration of 3% in this study, because we believe this is appropriate to achieve a sufficient antimicrobial effect and, based on the current reports, is not related with excessive itching and pain when used clinically. The effect of acetic acid in our study was compared to the bactericidal effect of commonly used topical agents.

1.

Materials and methods

The following Gram-negative and Gram-positive bacterial strains were tested: Escherichia coli (DSM Nr. 498, K12), Proteus vulgaris (Hauser 1885, group 2), Pseudomonas aeruginosa (Schroeter 1872, group 2), Acinetobacter baumannii (Institute Limbach, Heidelberg, Germany), Enterococcus faecalis (Andrews and Horder 1906, group 2), Stapylococcus epidermidis (Winslow 1908, group 2), MRSA (Institute Limbach, Heidelberg, Germany), Stapylococcus aureus (Rosenbach 1984, group 2) and b-

haemolytic Streptococcus group A and B (Institute Limbach, Heidelberg, Germany). All bacteria were isolated from patients in our burn unit. Standard 1-boullion-tubes were inoculated with these bacteria and incubated at 37 8C for 24 h. This is a standard method for the incubation of germs [2]. From the obtained bacterial cultures, 0.1 ml was mixed with 9.9 ml acetic acid (3%) and incubated for 5, 30 and 60 min each at a temperature of 37 8C. Dilutions of 10 2, 10 4 and 10 5 were prepared and covered with warm standard agar (47 8C). After incubation at 37 8C for 48 h, the colony-forming units (CFU) were counted. The testing of the other antiseptic agents was performed in the same way. The standards for performing bacterial cultures were based on the recommendations of the Robert Koch Institute and the German Institut of Hygiene [6]. The tested bactericidal solutions include povidone–iodine, 11% (Betaisodona1); polyhexanide, 0.04% (Lavasept1); mafenid acetate, 5% and chlorhexidine gluconate 1.5% cetrimide 15% (Hibicet1) and are routinely used in our department; an H2O group functioned as a control. Each combination of antiseptic agent and germ was tested four times and the mean CFUs were calculated for each possible combination.

2.

Results

Tables 1–3 summarise the results of the different antiseptic agents against the different germs; the calculated data are the mean of the four test results for each germ–antiseptic combination. After 5 min of incubation, there were no detectable CFUs of P. vulgaris, P. aeruginosa, A. baumannii and b-haemolytic Group B Streptococci in the acetic-acid-treated group (Tables 1 and 2). After 30 min, the growth of S. epidermidis (Table 2), S. aureus (Table 1) and b-haemolytic Group A Streptococci (Table 3) was completely inhibited at all dilutions. Escherichia coli, E. faecalis and MRSA (Table 3) were eliminated after 30 min at dilutions of 10 4 and 10 5; after 60 min, no plate showed countable bacterial growth after treatment with acetic acid. Most of the other tested agents showed complete elimination of all bacteria after 5 min, with the exception of P. vulgaris in a dilution of 10 2. Complete extinction was achieved after 30 min of incubation, in this case (Table 1). With A. baumannii in dilution of 10 2, only Hibicet1 took more than 5 min until no more CFUs could be observed (Table 1). In problematic Gram-negative bacteria, such as P. vulgaris, P. aeruginosa and A. baumannii, acetic acid showed better bacterial elimination after 5 min than any other antiseptic agent. Escherichia coli, E. faecalis, S. epidermidis and b-haemolytic Streptococcus group A were not eliminated after 5 min, but after 30 and 60 min each, respectively. The b-haemolytic Streptococcus group B has been eliminated equally by all antiseptics. P. vulgaris needed either a longer time for eradication by mafenid acetate, Lavasept1, Octenisept1 and Betaisodona1 or a greater dilution. Acetic acid and Hibicet1 seem to be more effective with this germ. In ‘problematic’ bacteria, such as P. vulgaris, P. aeruginosa and A. baumannii, acetic acid had a more effective inhibition

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burns 35 (2009) 695–700

Table 1 – Elimination of Proteus vulgaris, Pseudomonas aeruginosa and Acinetobacter baumannii by acetic acid 3%, povidone– iodine 11% (BetaisodonaW), polyhexanide 0,04% (LavaseptW), mafenid acetate 5% and chlorhexidine gluconate 1.5% cetrimide 15% (HibicetW) and H2O-control group. CFU: colony-forming units. Standard 1-boullion-tubes were inoculated with these bacteria and incubated at 37 8C for 24 h. From the obtained bacterial cultures, 0.1 ml was mixed with 9.9 ml acetic acid (3%) and incubated for 5, 30 and 60 min each at a temperature of 37 8C. Dilutions of 10S2, 10S4 and 10S5 were prepared and covered with warm standard agar (47 8C). After incubation at 37 8C for 48 h the colony-forming units (CFUs) were counted. The same process was carried out with the other antiseptics. Each combination of antiseptic agent and germ was tested four times and the mean CFUs were calculated for each possible combination. Antiseptic agent

Germ

Proteus vulgaris

Dilution Acetic acid 3%

Mafenide Acetate

Lavasept

Octenisept

Betaisodona

Hibicet

H2O

10 10 10

2

10 10 10

2

10 10 10

2

10 10 10

2

10 10 10

2

10 10 10

2

10 10 10

2

5 min

4 5

4 5

4 5

4 5

4 5

4 5

4 5

30 min

Pseudomonas aeruginosa 60 min

5 min

30 min

Acinetobacter baumannii

60 min

5 min

30 min

60 min

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

>10 3 69 2.5

>10 3 37.5 4

>10 3 36 –

>10 3 >10 3 133

>10 3 >10 3 154.5

>10 3 >10 3 149

>10 3 >10 3 69.5

>10 3 >10 3 62

>10 3 >10 3 23.75

86.5 – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

88.25 – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

472.25 – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

>10 3 >10 3 >10 3

>10 3 >10 3 >10 3

>10 3 64.5 26.25

>10 3 >10 3 >10 3

>10 3 >10 3 >10 3

>10 3 >10 3 216.75

>10 3 >10 3 >10 3

>10 3 >10 3 >10 3

>10 3 >10 3 73.25

Table 2 – Elimination of b-haemolytic Streptococcus group B, Staphylococcus epidermidis and Staphylococcus aureus by acetic acid 3%, povidone–iodine 11% (BetaisodonaW), polyhexanide 0.04% (LavaseptW), mafenid acetate 5% and chlorhexidine gluconate 1.5%/cetrimide 15% (HibicetW) and H2O-control group. CFUs: colony-forming units. Antiseptic agent

Germ Dilution

Acetic acid 3%

Mafenide Acetate

Lavasept

Octenisept

Betaisodona

Hibicet

H2O

10 10 10

2

10 10 10

2

10 10 10

2

10 10 10

2

10 10 10

2

10 10 10

2

10 10 10

2

4 5

4 5

4 5

4 5

4 5

4 5

4 5

b-Haemolytic Streptococci B 5 min

30 min

60 min

Staphylococcus epidermidis 5 min 3

30 min

60 min

Staphylococcus aureus 5 min 3

30 min

60 min

– – –

– – –

– – –

>10 26.75 4

– – –

– – –

>10 63.25 9

– – –

– – –

>10 3 48 2

>10 3 54 9.25

>10 3 38 4

>10 3 284.5 46.25

>10 3 281 48

>10 3 263.5 53.5

>10 3 >10 3 88

>10 3 >10 3 27.75

>10 3 >10 3 86.5

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

>10 3 >10 3 >10 3

>10 3 62.75 >10 3

>10 3 >10 3 24.25

>10 3 >10 3 >10 3

>10 3 >10 3 >10 3

>10 3 275.5 99.75

>10 3 >10 3 >10 3

>10 3 >10 3 >10 3

>10 3 >10 3 586.25

698

Table 3 – Elimination of Staphylococcus aureus and b-haemolytic Streptococcus group A, Escherichia coli, Enterococcus faecalis and Methicillin-resistant Staphylococcus aureus (MRSA) by acetic acid 3%, povidone–iodine 11% (BetaisodonaW), polyhexanide 0.04% (LavaseptW), mafenid acetate 5% and chlorhexidine gluconate 1.5%/cetrimide 15% (HibicetW) and H2O-control group. CFUs: colony-forming units. Antiseptic agent

Germ Dilution

Acetic acid 3%

Lavasept

Octenisept

Betaisodona

Hibicet

H2O

2

10 10 10

2

10 10 10

2

10 10 10

2

10 10 10

2

10 10 10

2

10 10 10

2

4 5

4 5

4 5

4 5

4 5

4 5

4 5

Escherichia coli

Enterococcus faecalis

5 min

30 min

60 min

5 min

30 min

60 min

5 min

30 min

>10 3 >10 3 882.75

– – –

– – –

>10 3 >10 3 88.25

47.25 – –

– – –

>10 3 584.75 69.25

– – –

>10 3 >10 3 908.5

>10 3 >10 3 685.25

>10 3 >10 3 705

>10 3 >10 3 182.25

>10 3 >10 3 124

>10 3 >10 3 148.5

>10 3 >10 3 92.25

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

>10 3 >10 3 >10 3

>10 3 >10 3 >10 3

>10 3 >10 3 875.25

60 min

Methicillin resistant Staphylococcus aureus (MRSA) 5 min

30 min

60 min

– – –

>10 3 273.25 67.25

– – –

– – –

>10 3 >10 3 104.5

>10 3 >10 3 121

>10 3 >10 3 92.75

>10 3 >10 3 112.25

>10 3 >10 3 78.5

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

– – –

>10 3 >10 3 >10 3

>10 3 >10 3 >10 3

>10 3 >10 3 224.75

>10 3 >10 3 >10 3

>10 3 >10 3 >10 3

>10 3 785.5 365

>10 3 >10 3 >10 3

>10 3 >10 3 >10 3

>10 3 >10 3 568.25

burns 35 (2009) 695–700

Mafenide Acetate

10 10 10

b-Haemolytic Streptococci A

burns 35 (2009) 695–700

Fig. 1 – Inhibiton of problematic bacteria such as (a) Proteus vulgaris, (b) Pseudomonas aeruginosa, and (c) Acinetobacter baumanii by acetic acid, mafenid acetate and H2O calculated by number of colony-forming units (CFUs) over time.

compared to mafenid acetate – which had been one of our standard antiseptics at this time – as shown in Fig. 1a–c. Because there was an overall lack of efficacy in the action of mafenide acetate against the different germs, we also performed resistance testing. Different degrees of resistance with erased mean inhibition concentrations for the germs (all germs are isolates from our patients) against mafenide acetate were evident.

3.

Discussion

It has been shown that mortality and morbidity in burn patients is highly correlated to wound infection [1]. Bacterial colonisation has been associated with a higher risk of infection and failure of skin grafts [3]. Hence, one important aim in burn treatment is the prevention and treatment of bacterial contamination of the burned or already-grafted areas. Nonetheless, the ideal treatment is still to be found. Nowadays, different agents are applied to burn wounds in an attempt to reduce the risk of septic complications developing in, and arising from, the wounds [1].

699

In times of increasing resistance rates against systemic and local antibiotics, local antiseptic agents have again become more important in wound management. Compounds such as silver nitrate, mafenide acetate, povidone–iodine or polyhexanide have been used in an attempt to reduce wound infection and sepsis [2]. In this study, the commonly used agents in the burn unit demonstrated excellent elimination of all tested germs, but have various known side effects. In 2003, Kalteis et al. showed in vivo tissue toxicity of many local antiseptics, which might cause prolonged wound healing [7,8]. Absorption might lead to severe metabolic problems. Steen reported several cases where application of povidone–iodine in extensive burns induced acidosis and even renal failure [9]. Apart from skin reactions such as itching or redness occasionally seen after treatment with local antiseptics, some cases of severe anaphylactic reactions to Lavasept1 have been described in the literature [6]. It is obvious that an alternative antiseptic agent, especially as a salvage choice in problematic bacteria, would be desirable in many clinical situations. Acetic acid has been commonly used in medicine for more than 6000 years for the disinfection of wounds and, especially, as an antiseptic agent in the treatment and prophylaxis of the plague. Earlier studies of Taylor [3], Phillips et al. [4] and Sloss et al. [5] are promising, but the data are very heterogeneous and account for small case numbers. The existing studies suggest effectiveness and clinical feasibility of acetic acid use with concentrations between 1% and 5%. Most effective elimination was achieved by soaking the wounds twice daily for 15 min; immersion baths proved to be less successful. No significant failures of skin grafts and no discomfort in the treated patients have been reported, but there were reports of odour and stinging sensation when the acid was used in concentrations higher than 5% [5]. The main goal of this study was to prove the suitability of acetic acid, in a low concentration of 3%, as a local antiseptic agent for the entire, typical spectrum of germs in an intensivecare burn unit. We used a 3% solution, because former reports suggested ideal antimicrobial activity between 1% and 5% and no reports about pain and itching could be found with 3% acetic acid. Excellent bactericidal effect of acetic acid was seen, particularly with problematic Gram-negative bacteria such as P. aeruginosa and A. baumannii. In P. vulgaris, acetic acid was even superior to commercially available antiseptics. P. vulgaris needed, for eradication, either a longer time of treatment by mafenidacetat, Lavasept1, Octenisept1 and Betaisodona1 or a greater dilution. Acetic acid and Hibicet1 seem to be more effective in this germ. This is an interesting study as it indicates that the time of exposure to Lavasept1, Octenisept1 and Betaisodona1 must be longer than 5 min to be effective, for example, for use in the treatment of post-skintransplantation surface contamination. Acetic acid or Hibicet1 could be used with P. vulgaris-positive swabs as an alternative. Because there was an overall lack in the efficacy of mafenid acetate against different germs, we also performed resistance testing. This showed different degrees of resistance with erased mean inhibition concentrations against mafenid acetate, indicating that selection of our ‘clinic-specific germs’ has occurred over time. This is possible because mafenide

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acetate has a sulfonamide-group, like antibiotics of the sulfonamide-type, which is a well-known target for resistance development. At present, mafenide acetate or other local antiseptics are locally applied for a 20-min application daily in patients with positive-swab results for problematic germs such as P. vulgaris, P. aeruginosa and A. baumannii. If this is application proves ineffective after several days, based on our own data, we must switch to another local antiseptic such as acetic acid. Focussing on these germs, acetic acid had a more effective bacterial inhibition compared to mafenide acetate; therefore, acetic acid might be an alternative for use against these problematic germs. The exact mechanism of acetic acid is unknown, but a physical alteration of the bacterial cell wall is presumed. This might be the reason for its higher effectiveness in Gram-negative, than in Gram-positive, bacteria because of the less resistant cell wall in Gram-negative bacteria caused by the lack of murein. The microbiological spectrum is wide, even at the tested low concentration of 3%. Compared to our currently used antiseptic solutions, it showed, in some bacteria, similar or even better bactericidal properties. At the concentration of 3%, itching and pain would not appear. Although there have been earlier studies investigating the bactericidal effect of acetic acid, this study is the first to compare the bactericidal effect of commonly used topical antiseptics with the effect of acetic acid in a wide spectrum of pathological bacteria found in a burn unit. Earlier studies have focussed on singular problematic germs such as P. aeruginosa, but the spectrum at an intensive-care burn unit is quite different. Based on the promising results of this investigation and the already proven suitability for the application in burn wounds in a pilot study, a clinical study is currently underway to evaluate the efficacy of acetic acid in the reduction of burnwound contamination. We have already performed successful individual healing therapy with acetic acid in four patients with multi-drug-resistant P. aeruginosa or multi-drug-resistant A. baumannii infections as a salvage procedure. In these cases, after a daily treatment with acetic acid for 20 min, the follow-

up swab diagnostics were reported negative. It is yet to be proven whether natural acetic acid has significant benefits with regards to wound healing and bacterial growth compared to industrial acetic acid. It can be concluded from the data that acetic acid in a concentration of 3% could be suitable as a local antiseptic agent.

Conflict of interest statement There is no conflict of interest.

references

[1] Brown TP, Cancio LC, McManus AT, Mason Jr AD. Survival benefit conferred by topical antimicrobial preparations in burn patients: a historical perspective. J Trauma 2004;56(4):863–6. [2] Willenegger H. Local antiseptics in surgery—rebirth and advances. Unfallchirurgie 1994;20(2):94–110. [3] Taylor K. Treatment of Bacillus pyocyaneus infection. JAMA 1916;67:1598–9. [4] Phillips I, Lobo AZ, Fernandez R, Gundara NS. Acetic acid in the treatment of superficial wounds infected by Pseudomonas aerigunosa. Lancet 1968;i:11–3. [5] Sloss JM, Cumberland N, Milner SM. Acetic acid used for the elimination of Pseudomonas aeruginosa from burn and soft tissue wounds. J R Army Med Corps 1993 Jun;139(2):49–51. [6] Liste der vom Robert Koch Institut gepru¨ften und anerkannten Desinfektionsmittel und –verfahren. Bundesgesundheitsblatt 2003: (46); 72–95. [7] Gilliland EL, Nathwani N, Dore CJ, Lewis JD. Bacterial colonisation of leg ulcers and its effect on the success rate of skin grafting. Ann R Coll Surg Engl 1988 Mar;70(2):105–8. [8] Kalteis T, Luring C, Schaumburger J, Perlick L, Bathis H, Grifka J. Tissue toxicity of antiseptics. Z Orthop Ihre Grenzgeb 2003;141(2):233–8. [9] Steen M. Review of the use of povidone–iodine (PVP–I) in treatment of burns. Postgrad Med J 1993;69(Suppl. 3):S84–92.