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Anti-microbial Properties of EpiThera®

Technical Bulletin April 2004 TB10-04

Introduction

EpiThera® is a cost-effective, easily applied topical spray that promotes healing of surgical incisions, mechanical wounds and abrasions, and burns. Wound healing is accelerated, and the ultimate appearance of the wound site is such that scar tissue formation is greatly reduced.
One of the important attributes of EpiThera® is its apparent anti-microbial properties, based on testimonials that show a very low incidence of microbial infection. The purpose of this study was to assess the anti-microbial properties of EpiThera® in a controlled, laboratory setting.

Pathogenic Microbes Tested

The following microbes were tested in this study:

These bacteria are common pathogens.

Methodology Employed

A standard way of assessing anti-microbial properties is to introduce the material being tested into a liquid suspension of bacteria in a nutrient broth. (Whichard JM, Sriranganathan N, Pierson FW, 2003) As the bacterial population in the suspension grows, the turbidity (optical density) of the suspension will increase. The greater the turbidity, the higher the bacterial population. The introduction of an anti-microbial agent that kills bacteria will reduce the turbidity (optical density) of the solution. This technique provides a fast and convenient way of testing the anti-microbial properties of liquid formulations such as EpiThera®.


Anti-microbial tests of EpiThera® were carried out in the laboratory of Dr. Daniel Kunz, Department of Biological Sciences, University of North Texas, Denton, Texas. All bacteria were grown as shelf cultures at 37oC. The sole exception of C. albicans which was grown at 30oC. All bacteria were grown in Mueller-Hinton (M-H) medium, which is the standard medium used for anti-microbial susceptibility testing.


Aliquots of EpiThera® were added to sterile diluted liquid culture medium to a total end volume of 5 ml (see below). All tubes were then inoculated with 0.1 ml of an overnight broth culture (~1-2 X 106 cells [CFU]). Bacterial growth was determined after 24 h by determining culture turbidity using a spectrophotometer to determine optical density at 540 nm.
Anti-microbial properties were assessed using the following concentrations of EpiThera® in medium: 0%, 0.1%, 0.2%, 0.5%, 1%, 2% and 5%. Reference blanks contained sterile medium with EpiThera® at each of the above concentrations, but without inoculated cells.


Data were expressed as the minimal inhibitory concentration (MIC), which is defined as the lowest concentration of EpiThera® inhibiting all microbial growth as evidenced by a lack of detectable turbidity in the medium.

Findings of the Study

EpiThera® showed anti-microbial properties against all eight microbes tested. Figure 1 shows a plot of optical density of the medium (which is directly proportional to cell concentration in the medium) against concentration of EpiThera® in the medium.

Figure 1. Medium Optical Density as a function
of EpiThera® concentration.
emu-oil-epithera

Table1. Minimal Inhibitory Concentrations of EpiThera® in eight microorganisms.

microorganism

As evident from the sharp fall in optical density as EpiThera® concentration increased(Figure 1), EpiThera® began to suppress microbial growth at concentrations as little as 0.5% in most microbes tested. At 2% concentration in the medium, EpiThera® completely suppressed the growth of the bacteria Escherichia coli, Salmonella typhimurium, Pseudomonas aeruginosa, and Enterococcus faecalis.


A 5% concentration suppressed growth of the bacteria Staphylococcus aureus, Bacillus cereus, Streptococcus agalactiae and the yeast Candida albicans (Table 1).

Interpretations and Conclusions

In vitro EpiThera® shows clear suppression of the growth of bacteria and yeast when in aqueous solution. While minimal inhibitory concentrations (MIC) vary, all MICs were 5% or less for the eight microbes tested, and most showed some significant decrease in culture growth rate at concentrations of EpiThera® of as little as 0.1% (e.g. Salmonella typhimuriu, Escherichia coli).
Somewhat confounding strict interpretation of these type data is that in vitro testing occurred by diluting EpiThera® in the test medium, a typical laboratory procedure for in vitro testing of anti-microbial properties. How these in vitro data relate to the actual concentration of EpiThera® occurring when the formulation is topically applied to a wound as directed has not been determined. However, since EpiThera® isapplied at full strength (100% concentration) in human applications, the concentration of EpiThera® experienced at least by pathogens on the surface tissues of wounds is likely to be high, and certainly above the MICs reported in this study.


It is concluded that EpiThera® has significant anti-microbial properties in vitro in a wide range of microbial pathogens. Furthermore, EpiThera® is very likely to highly suppress microbial pathogens located superficially in fresh or recovering wounds.

References

Whichard JM, Sriranganathan N, Pierson FW 2003.
Suppression of Salmonella growth by wild- type and large-plaque variants of bacteriophage Felix 01 in liquid culture and on chicken frankfurters. J Food Prot. 66:220-5.

EpiThera® is a registered trademark of Lucid Med Tec II, Ltd.