Encyclopedia of Environmental Science and Engineering, Volume I and II

DISINFECTION 229

the hypothesis that disinfectants in use shall be at least as

efficient as 5% phenol, and that a dilution of twenty times

the phenol coefficient should achieve this. Accordingly, this

dilution is tested against standardized cultures in order to

confirm this, or alternatively, in order to determine a correc-

tion factor.

Whereas the extinction methods discussed above evaluate

the extinction concentration corresponding to predetermined

exposure times, Berry and Bean (1954) devised a test for

evaluating extinction times for chosen disinfectant concentra-

tions. While the test is only applicable to phenolic and other

easily inactivated disinfectants, it has been claimed to be at

least as reproducible as other methods in common use (Cook

and Wills, 1954). The method of assessing the end-point of

the reaction has been further improved by Mathei (1949).

Methods other than extinction tests for standardizing dis-

infectants include various methods based on assessment of

cessation of vital enzyme activities. The enzymes involved

have generally been certain oxidases or dehydrogenases

(Sykes, 1939; Knox et al. , 1949). In a less specific manner,

inhibition of respiration has been used as a method of assess-

ment (Roberts and Rahn, 1946). The controversy surround-

ing these methods centers around the problem of correlating

enzyme activity with viability.

Other methods which have been proposed include mea-

surement of post-incubation opacity corresponding to stan-

dard survivor levels (Needham, 1947), and also measurement

of cell volume increase following post-exposure incubation

(Mandels and Darby, 1953).

The range of tests discussed above is by no means

exhaustive, and only covers general purpose tests appli-

cable to water-misible disinfectants. Any number of alter-

native tests could be devised by appropriate selection and

standardization of the various parameters. In addition, there

are numerous tests which can be, and have been, devised in

order to standardize disinfectants intended for specific uses

such as sporicidal or tuberculocidal duties (AOAC 1970).

Methods of standardizing disinfectants other than those

to which the above discussion applies are usually designed

more closely around the particular use envisaged. Thus, a

considerable element of actual, or simulated, in-use testing

is usually involved. For convenience, they are best discussed

in the following section.

A recent test is the Kelsey–Sykes test (Kelsey and Maurer,

1974), which is a form of capacity test. In this, incremental

additions of test organisms are made to appropriate dilutions

of test disinfectant and aliquots are removed for detecting

survival immediately prior to the next addition of organisms.

On the basis of this method, use-dilutions of the test dis-

infectant (which need not necessarily be a phenolic) under

clean and dirty conditions can be recommended to hospitals,

which should then check them during in-use tests (see also

Coates, 1977; Cowen, 1978).

In-Use Tests

The previous two sections have dealt with testing methods

applied in the laboratory, which yield information primarily

of value to the disinfectant manufacturer. The “consumer,”

however, is almost solely concerned with the performance

of disinfectant materials under the conditions of use which

are associated with the application envisaged. To this end,

he is more interested in testing methods which resemble, as

closely as possible, these practical conditions. However, the

foregoing remarks should not be taken to imply that labo-

ratory standardization methods are completely arbitrary. In

view of the greater difficulty experienced in killing organ-

isms which are present as dried surface films, as opposed to

those in fluid suspension, many standardization tests have

employed films on various surfaces as their inocula. Surfaces

used have included silk thread (Koch, 1881), garnets (Kronig

and Paul, 1887), glass cover-slips (Jensen and Jensen, 1933),

glass cylinders (Mallmann and Hanes, 1945), glass slides

(Johns, 1946), stainless steel cylinders (AOAC Use-Dilution

Confirmatory Test, 1960), and glass tablet tubes (Hare, Raik

and Gash, 1963). It should be noted that while these sur-

faces represent a step toward the practical situation, they

nevertheless comprise a collection of laboratory “artifacts”

when compared with real situations. A nearer approach was

achieved by use of surfaces such as rubber strips (Goetchins

and Botwright, 1950) and glazed, waxed, and rubber tiles

(Rogers, Mather and Kaplan, 1961).

Where the physical size of articles required to be disin-

fected is fairly small, it is quite feasible to carry out in-use

testing in the laboratory. Hence metal trays were used by

Neave and Hoy (1947), 10 gallon milk churns by Hoy and

Clegg (1953), and small drinking glasses by Gilcreas and

O’Brien (1941). Similarly, scalpels, syringes and similar

small items may conveniently be subjected to in-use testing

in the laboratory. Where the physical size of the system is

somewhat greater, then the laboratory must be forsaken in

order to carry out on-site testing. Typical examples of such

situations include hospital walls and floors, and industrial or

dairy processing machinery. The outstanding value of in-use

testing arises from the fact that results obtained are directly

applicable to the system without the need for interpreta-

tion and extrapolation, and that practical difficulties such as

short contact times or inaccessibility of certain areas can be

accounted for.

The choice of test organisms usually reflects either of

two main approaches. The simplest approach is to inoculate

the system artificially with organisms considered to be of

practical significance in the particular application. This sig-

nificance may be due to the resistance to disinfection of the

organism, or alternatively, to its practical effect on the system

should it survive the disinfection process. In order to simu-

late the practical system, the organisms may be suspended

in appropriate materials before inoculation. An example

would be the use of milk as a suspending fluid in tests on

dairy disinfection. A slight variation on this approach is to

inoculate with indeterminate mixtures of likely organisms

obtained from natural sources, such as low quality, raw milk.

The second approach involves the use of the normal, pre-

existing flora of the system which has arisen during normal

use. Investigations of this flora before and after the disin-

fection process would provide direct evidence as to whether

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