The Minimum Inhibitory Concentration (MIC) is defined as the lowest concentration of an antimicrobial ingredient or agent that is bacteriostatic (prevents the visible growth of bacteria). MICs are used to evaluate the antimicrobial efficacy of various compounds by measuring the effect of decreasing concentrations of antibiotic/antiseptic over a defined period in terms of inhibition of microbial population growth. These evaluations can be quite useful during the R&D phase of a product to determine appropriate concentrations required in the final product, as the concentration of drug required to produce the effect is normally several hundred to thousands of times less than the concentration found in the finished dosage form.
Various concentrations of the compounds are inoculated with cultured bacteria, and the results are measured using agar dilution or broth dilution (macro or micro) to determine at what level the MIC endpoint is established. Susceptibility testing is typically conducted using organisms that contribute to an infectious process warranting antimicrobial chemotherapy. A commonly used cocktail of bacteria is known as the ESKAPE pathogens (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter species). The ESKAPE pathogens are considered the leading cause of nosocomial (hospital-related) infections and are known to be resistant to antimicrobial products. Other organisms such as aerobic or anaerobic bacteria, yeasts, or filamentous fungi, either alone or in multiple combinations can also be utilized. The FDA Tentative Final Monograph (TFM- section 333.470, page 31444) also provides guidance on organisms to use to determine the efficacy of products such as antiseptic handwash, healthcare personnel handwash, patient preoperative skin preparations, and surgical hand scrubs.
The Minimum Bactericidal Concentration (MBC) is the lowest concentration of an antibacterial agent required to kill a bacterium over a fixed, somewhat extended period, such as 18 hours or 24 hours, under a specific set of conditions. It can be determined from the broth dilution of MIC tests by subculturing to agar plates that do not contain the test agent. The MBC is identified by determining the lowest concentration of antibacterial agent that reduces the viability of the initial bacterial inoculum by a pre-determined reduction such as ≥99.9%. The MBC is complementary to the MIC; whereas the MIC test demonstrates the lowest level of antimicrobial agent that greatly inhibits growth, the MBC demonstrates the lowest level of antimicrobial agent resulting in microbial death. In other words, if a MIC shows inhibition, plating the bacteria onto agar might still result in organism proliferation because the antimicrobial did not cause death. Antibacterial agents are usually regarded as bactericidal if the MBC is no more than four times the MIC. MBC testing can be a good and relatively inexpensive tool to simultaneously evaluate multiple antimicrobial agents for potency. 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 Clinical and Laboratory Standards Institute (CLSI) has established protocols and standards for establishing MIC and MBC in products. A common methodology utilized for MIC is CLSI M07-A9, Methods for Dilution Antimicrobial Susceptibility Tests for Bacteria That Grow Aerobically. CLSI also has developed methods specific for the yeasts, filamentous fungi, and anaerobic bacteria. For MBC determination, CLSI M26-A, Methods for Determining Bactericidal Activity of Antimicrobial Agents, is an accepted industry standard.