Control by Low Temperature ◾ 461
freezes below 0°C). As the temperature drops further and more ice crystals form, the solutes get
concentrated in the remaining water, which, in turn, depresses the freezing point of the water in
the solution. The A W is also reduced. When the temperature is reduced to approximately –20°C
(–4°F), almost all the free water freezes.
As the temperature of a food is reduced below –2°C (28.4°F), free water inside the microbial
cells also undergoes similar changes. At a slow rate of freezing, as the water molecules in the food
start freezing, water molecules from inside the microbial cells migrate outside, causing dehydration of cells and concentration of solutes and ions inside. When the temperature is reduced further
(below –20°C) so that the water in the food has frozen, water inside the cell also freezes. However,
before that, microbial cells are exposed to low pH (resulting from concentration of ions) and low
A W (resulting from concentration of solutes) inside and outside the cells. This can cause denaturation and destabilization of the structural and functional macromolecules (enzymes) in the
microbial cells, whose stability and functions depend on their three-dimensional structures, and
can injure the cells. If the freezing is rapid, very small ice crystals form quickly, and the cells are
not exposed to the solution effect. This is practiced in the freeze preservation of starter cultures
and frozen stock cultures.
Microbial cells subjected to freezing and thawing suffer sublethal (repairable) as well as lethal
injury. Studies show that different components of the cell wall (or outer membrane) and cell membrane (or inner membrane) are injured. DNA strand break, ribosomal RNA degradation, and activation and inactivation of some enzymes have also been reported in some studies. In sublethally
injured cells, the structural and functional injuries are reversible. In lethally injured (or dead) cells,
the damages are irreversible (Chapter 10). 2–4
influencing Factors
The effectiveness of low temperatures in controlling microbial growth and microbial enzymatic
activity in food depends on many factors. These factors can be arranged into three groups: those
unique to low temperatures, those related to the food environment, and those inherent in microorganisms. These factors not only help prevent or reduce the growth of microorganisms, but can
also greatly influence the extent of sublethal and lethal injury that microorganisms incur in food
preserved at low temperatures. An understanding of the influence of these factors and interaction
among them is important to design an efficient and predictable method to preserve a specific food
at low temperature.
Nature of Process
At temperatures above freezing of free water (≤–2°C), different types of bacteria, molds, and
yeasts can grow in a food. But the lag and exponential phases become longer as the temperature
is reduced. In the low range, even a difference in <1°C can be highly important. A Pseudomonas
fluorescens strain was reported to have a generation time of approximately 6.7 hours at 0.5°C
but 32.2 hours at 0°C. Thus, a reduction in 0.5°C increased the generation time by approximately 4.5-fold. This is much more than the theoretical estimate, which suggests that the
generation time doubles for every 10°C (50°F) reduction. Spores of some spoilage Bacillus and
Clostridium spp. can germinate even at refrigeration temperatures (4.5°C or 40°F). Cells of
some mesophiles and thermophiles can be sublethally injured as well as die as the temperature
drops below 4.5°C.
freezes below 0°C). As the temperature drops further and more ice crystals form, the solutes get
concentrated in the remaining water, which, in turn, depresses the freezing point of the water in
the solution. The A W is also reduced. When the temperature is reduced to approximately –20°C
(–4°F), almost all the free water freezes.
As the temperature of a food is reduced below –2°C (28.4°F), free water inside the microbial
cells also undergoes similar changes. At a slow rate of freezing, as the water molecules in the food
start freezing, water molecules from inside the microbial cells migrate outside, causing dehydration of cells and concentration of solutes and ions inside. When the temperature is reduced further
(below –20°C) so that the water in the food has frozen, water inside the cell also freezes. However,
before that, microbial cells are exposed to low pH (resulting from concentration of ions) and low
A W (resulting from concentration of solutes) inside and outside the cells. This can cause denaturation and destabilization of the structural and functional macromolecules (enzymes) in the
microbial cells, whose stability and functions depend on their three-dimensional structures, and
can injure the cells. If the freezing is rapid, very small ice crystals form quickly, and the cells are
not exposed to the solution effect. This is practiced in the freeze preservation of starter cultures
and frozen stock cultures.
Microbial cells subjected to freezing and thawing suffer sublethal (repairable) as well as lethal
injury. Studies show that different components of the cell wall (or outer membrane) and cell membrane (or inner membrane) are injured. DNA strand break, ribosomal RNA degradation, and activation and inactivation of some enzymes have also been reported in some studies. In sublethally
injured cells, the structural and functional injuries are reversible. In lethally injured (or dead) cells,
the damages are irreversible (Chapter 10). 2–4
influencing Factors
The effectiveness of low temperatures in controlling microbial growth and microbial enzymatic
activity in food depends on many factors. These factors can be arranged into three groups: those
unique to low temperatures, those related to the food environment, and those inherent in microorganisms. These factors not only help prevent or reduce the growth of microorganisms, but can
also greatly influence the extent of sublethal and lethal injury that microorganisms incur in food
preserved at low temperatures. An understanding of the influence of these factors and interaction
among them is important to design an efficient and predictable method to preserve a specific food
at low temperature.
Nature of Process
At temperatures above freezing of free water (≤–2°C), different types of bacteria, molds, and
yeasts can grow in a food. But the lag and exponential phases become longer as the temperature
is reduced. In the low range, even a difference in <1°C can be highly important. A Pseudomonas
fluorescens strain was reported to have a generation time of approximately 6.7 hours at 0.5°C
but 32.2 hours at 0°C. Thus, a reduction in 0.5°C increased the generation time by approximately 4.5-fold. This is much more than the theoretical estimate, which suggests that the
generation time doubles for every 10°C (50°F) reduction. Spores of some spoilage Bacillus and
Clostridium spp. can germinate even at refrigeration temperatures (4.5°C or 40°F). Cells of
some mesophiles and thermophiles can be sublethally injured as well as die as the temperature
drops below 4.5°C.