Rate of kill

When bacterial cells are shifted directly from a low growth temperature to a lethal temperature, the cells are rapidly killed. However, if the cells are first preadapted by growth at a non-lethal temperature for 30 min, the rate of killing upon a shift to lethal temperature is dramatically decreased these cells have acquired thermotolerance. During the pre-adaptation phase a heat shock response is induced which leads to increased synthesis of heat shock proteins. 

Inspection of the death curves obtained from viable count data had early ehcited the idea that because there was usually an approximate, and under some circumstances a quite excellent, linear relationship between the logarithm of the number of survivors and time, then the disinfection process was comparable to a unimolecular reaction. This imphed that the rate of killing was a function of the amount of one of the participants in the reaction only, i.e. in the case of the disinfection process the number of viable cells. From this observation there followed the notion that the principles of first-order... 

The more usual pattern found experimentally is that shown by B, which is called a sigmoid curve. Here the graph is indicative of a slow initial rate of kill, followed by a faster, approximately linear rate of kill where there is some adherence to first-order reaction kinetics this is followed again by a slower rate of kill. This behaviour is compatible with the idea of a population of bacteria which contains a portion of susceptible members which die quite rapidly, an aliquot of average resistance, and a residue of more resistant members which die at a slower rate. When high concentrations of disinfectant are used, i.e. when the rate of death is rapid, a curve ofthe type shown by C is obtained here the bacteria are dying more quickly than predicted by first-order kinetics and the rate constant diminishes in value continuously during the disinfection process. 

A, obtained if the disinfection process obeyed the first-order kinetic law. B, sigmoid curve. This shows a slow initial rate of kill, a steady rate and finally a slower rate of kill. This is the form of curve most usually encountered. C, obtained if bacteria are dying more quickly than first-order kinetics would predict. The constant, K, diminishes in value continuously during the process. 

The similarity between Equations 1 (mite) and 2 (egg) is remarkable considering each was derived from independent biological datasets this result is consistent with the concept, verified by independent experiments (12) on rate of kill vs. egg age, that egg kill with this series of compounds involves kill of pharate larvae rather than prevention of emergence. 

The D value is a single quantitative expression of the rate of killing of microorganisms. The D term refers to the decimal point in which microbial death rates become positive time values by determining the time required to reduce the microbial population by one decimal point. This is also the time required for a 90% reduction in the microbial population. Hence, the time or dose it takes to reduce 1000 microbial cells to 100 cells is the D value. The D value is important in the validation of sterilization processes for several reasons. 

For multidose sterile products, the preservative must be capable of reestablishing sterility between each use, whereas for a non-sterile topical cosmetic the function of the preservative might simply be to prevent growth. The associated toxicity of preservatives often limits the concentrations at which they can be employed thus, lower concentrations are generally employed for opthal-mic products and injectables. In choosing a preservative the likely capacity required, the rate of killing desired, and the ingredients and pH of the formulation must be borne in mind. 

In practice, such obedience to first-order kinetics is rare for chemical inactivation. More commonly, deviations from first-order kinetics are obtained, when there is either an initial lag in the rate of killing (Fig. 2A) or when the rate of killing decreases with time of exposure (Fig. 2B). The former is commonly observed when the concentrations of preservative are... 

Fig. 2 Deviations from first-order kinetics of the inactivation of microbial populations by treatment with chemical agents due to (A) an initial lag in the rate of killing or (B) a decrease in the rate of killing with time of exposure.

Approximation to first-order kinetics implies that the sensitivity toward preservatives of a population of microorganisms is normally distributed. Decreases in the rate of killing with time often results from adsorptive loss of thepreservative onto and into the killed cells, causing a decrease in the available concentration of the preservative for killing the surviving organisms. The susceptibility of preservatives to such adsorptive losses is described as its capacity. 

The rate of killing of an antimicrobial agent is directly dependent on the temperature of the interaction. This dependence is described by the temperature coefficient Q or 0), which can be determined either from the slope of the graph relating the Z)-value or K and temperature, or from Eq. (6). 

When preservatives are added to two-phase oil-water systems, depending upon the hydrophilicity of the agent, they partition between the two phases. Bacteria will only grow within the aqueous phase or at the oil-water interface. Thus, preservative lost to the oil is unavailable for antibacterial action. Hence the concentration in the water phase is reduced, affecting the rate of kill, but the capacity of the system is unaffected as... 

Hurwitz SJ, McCarthy TJ. The effect of pH and concentration on the rates of kill of benzoic acid solutions against E. coli. Clin Pharm Ther 1987 12 107-115. 

It is also possible to plot the rate of kill against the temperature. 

The properties of an ideal preservative are well recognized a broad spectrum of activity and a rapid rate of kill selectivity in reacting with the contaminants and not the formulation ingredients nonirritant and non-toxic to the patient and stable and effective throughout the life of the product. 

Most preservatives interact in solution to some extent with many of the commonly used formulation ingredients via a number of weak bonding attractions as well as with any contaminants present. Unstable equilibria may form in which only a small proportion of total preservative present is available to inactivate the relatively small microbial mass the resulting rate of kill may be far lower than might be anticipated from the performance of simple aqueous solutions. However, unavailable preservative may still contribute to the general irritancy of the product. It is commonly believed that where the solute concentrations are very high, and Aw is appreciably reduced, the efficiency of preservatives is often significantly reduced and they may be virtually inactive at very low Aw. The practice of in-... 

Isquith, A. ]. and McCollum, C. J. "Surface kinetic test method for determining rate of kill by an antimicrobial solid Appl. Environ. Microbiol. 1978. 36 700-704. 

FIGURE 14.22. The effect of //on rate of kill of Escherichia coli with glutaraldehyde... 

The effect of concentration on a mixed culture at different pH was demonstrated by Grab et a/[1990] and is shown on Fig. 14.23. a combination of pH 8.1 and a glutaraldehyde concentration of 200 mg/l gave a remarkable rate of kill, i.e. reducing viable cell numbers from around 5 x 10 to near zero in about 1 hour. 

Also required is a study to assess whether the antiseptic is fast-acting, as described by the definition for an indication (Table 4). The time-kill kinetic study is designed to measure the rate of kill by the antiseptic under controlled conditions. The time-kill kinetic study is performed with a select group of gram-... 

Another characteristic desirable in antiseptics used by healthcare professionals is that they act as microbiologically lethal agents. The spectmm of activity study does not allow evaluation of this characteristic since incubation exceeds 18 hours. However, the time-kill kinetic study does allow assessment of the rate of kill of an antiseptic provided certain variables are controlled [34]. 

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