Esterilizacion

Microbioogical

Process Discussion

Calculation of Heat Sterilization Times for Fermentation Media
FRED H. DEINDOERFER1
Merck Sharp & Dohme Research Laboratories, Merck & Co., Inc., Rahway,

New

Jersey

Received for publication September 14, 1956

Sterilization of nutrient media is an operation
essential to all industrial fermentation processes requiring pure culture maintenance.AMethods for sterilizing nutrient media include 1) heating, 2) filtration,
3) irradiations, 4) sonic vibration, and 5) exposure to
chemical agents. Because heating the medium, usually
by steam, is the most reliable and on a large scale the
easiest to control, it is the method of choice throughout
the fermentation industry. The increasing availability
of radioactive isotopes undoubtedly willstimulate
further study of the use of gamma radiation for sterilization. However, a great deal of work is needed before
commerically feasible gamma ray applications will be
used in the fermentation industry. The other methods,
although successful on a laboratory scale, present too
many operational drawbacks for large-scale use in
fermentation processes.
How long should a nutrient medium beexposed at
high temperatures to achieve sterile conditions? This
question arises often and usually is answered in benchscale or pilot plant tests. Improper translation of these
test results with large equipment may subject the
medium to unnecessary overheating. A method by
which minimum exposure time to achieve sterile conditions can be calculated from easily obtainable thermaldeathrelationships and the temperature conditions in a
heat sterilization process is presented in this paper. The
method can be used to correlate sterilization conditions
among various sized fermentation vessels. It also permits evaluation of the temperature and retention time
relationship in continuous sterilizers. It need not be
restricted to fermentation processes, but should be
applicable to anyprocess involving a heat sterilization
or pasteurization operation.

Bacterial spores are by far the most heat resistant forms
of microorganisms.
The thermal resistance of bacterial spores is inherently different among species and even among strains
of the same species. Besides thermal resistance inherent
to spores of a particular species, variations can be caused
by a number of environmentalfactors. These can be
generalized into two groups: 1) environmental factors
affecting sporulation prior to sterilization, and 2)
environmental factors affecting spores during the
sterilizing heat exposure. The most important factors
in nutrient media sterilization fall in the latter category
and include factors such as the pH during sterilization
and the osmotic nature of the media. Thepresence of
suspended solids also affects sterilization by physically
insulating spores from heat exposure.
Thermal-Death Relationships of Bacterial Spores
Because bacterial spores are the most heat resistant
forms of microorganisms, their germinated cells are the
most frequent contaminants encountered in industrial
fermentations due to improper sterilization. The ensuing
discussion will deal,therefore, with the thermal-death
relationships of these forms. Relationships between the
number of viable spores and exposure time to heat
demonstrate a logarithmic rate of spore viability destruction. Two typical survivor curves for spores of
Bacillus stearothermophilus strain 1518, an organism
often used for sterilization studies in the food industry,
are shown in figures 1A and lB.These curves were
obtained in buffer solutions and would differ from their
respective curves in other media. Spore destruction at
the higher temperature is more than 400 times faster
than at the lower temperature.

Thermal Resistance of Microorganisms
The ability of microorganisms to withstand heat is
much greater than that of other forms of life. Thermophiles capable of even tolerating...