The MBC test determines the lowest concentration at which an antimicrobial agent will kill a particular microorganism. The MBC is determined using a series of steps, undertaken after a Minimum Inhibitory Concentration (MIC) test has been completed.
MBC testing is useful for comparing the germ-killing activity of several antimicrobial agents at once.
Summary of the Minimum Bactericidal Concentration Test
- A pure culture of a specified microorganism grown overnight, then diluted in growth-supporting broth (typically Mueller Hinton Broth) to a concentration between 1 x 10^5 and 1 x 10^6 cfu/ml.
- A stock dilution of the antimicrobial test substance is made at approximately 100 times the expected MIC (if known).
- Further 1:1 dilutions are made in test tubes or 96 well microtiter plates.
- All dilutions of the test product(s) are inoculated with equal volumes of the specified microorganism.
- A positive and negative control tube or well is included for every test microorganism to demonstrate adequate microbial growth over the course of the incubation period and media sterility, respectively.
- An aliquot of the positive control is plated and used to establish a baseline concentration of the microorganism used.
- The tubes or microtiter plates are then incubated at the appropriate temperature and duration.
- Turbidity indicates growth of the microorganism and the MIC is the lowest concentration where no growth is visually observed.
- To determine the MBC, the dilution representing the MIC and at least two of the more concentrated test product dilutions are plated and enumerated to determine viable CFU/ml.
- The MBC is the lowest concentration that demonstrates a pre-determined reduction (such as 99.9%) in CFU/ml when compared to the MIC dilution.
Strengths of the Minimum Bactericidal Concentration Test
- The MBC test allows determination of the minimum concentration of an agent necessary to achieve a bactericidal effect. It is worth noting, however, that the duration of time the antimicrobial is in contact with the test organism is quite long for this method, on the order of 18 hours. Thus, the test truly does determine the minimum concentration needed to kill the test organism, since all other parameters are conducive to biocidal effect.
- The MBC test can be a good and relatively inexpensive tool to rank a great number of antimicrobial agents by potency, for screening purposes.
- The MBC test can be used to evaluate formulation problems wherein the formulator suspects that the active ingredient is being "bound up" by other ingredients. The theory is that the MBC will be worse for a formula that has a portion of its active ingredient chemically combined with other ingredients, thus not available to kill microorganisms in the suspension.
- The test parameters for the MBC are easy to control in the laboratory, so comparisons can be made fairly easily between various antimicrobial agents tested under the same conditions and their respective effects on specific microorganisms.
Weaknesses of the Minimum Bactericidal Concentration Test
- An MBC must be determined for each microorganism individually as the antimicrobial will likely have different MBC values for different test microorganisms.
- MBC testing results will not determine the concentration of the antimicrobial needed to disinfect or sanitize microorganisms at a short contact time, such as 10 minutes.
- More nutritive growth media such as Tryptic Soy Broth can negatively affect MBC values. Mueller Hinton Broth is the recommended broth for this method.
Microchem Laboratory routinely runs minimum bactericidal concentration (MBC) tests for its customers. The laboratory is well versed in the science of antimicrobial testing in general, and applies this knowledge to each and every study we undertake.
If you are interested in MBC testing, you are likely in the research and development rather than claim substantiation phase. Microchem Laboratory has an excellent track record of helping companies move from screening to GLP product testing.