Food Spoilage by Microbial Enzymes ◾ 289
raw milk, meat, fish, and other food products. During refrigerated storage, psychrotrophs are able
to grow and produce heat-stable enzymes in foods. 6,7
Production of heat-stable proteinases and lipases in milk by some of these bacteria, especially
Pseudomonas spp., and the characteristics of these enzymes have been well studied. The results show
that different species and strains produce proteinases and lipases that differ in molecular weight and
activity. Thus, a highly active proteinase or lipase produced by a Pseudomonas strain can produce
extensive proteolysis or lipolysis, even at 10 5–6 cells/mL milk. But another strain may need to reach
to 10 9 cells/mL to produce similar changes. The enzymes can be produced in raw milk at refrigerated temperatures (1°C–7°C) in sufficient quantities to hydrolyze proteins and lipids in detectable
levels within 3–7 days. The catalytic activity of the proteinases is highest between pH 6.0 and 7.0
with a pH range between 5.0 and 9.0. Pasteurization of milk (at 63°C for 30 minutes or 71°C for
15 seconds) results in a loss of 6%–36% activity and, even after heating at 121°C for 10 minutes,
some activity of the proteinases is retained. UHT treatment (140°C–150°C for 1–5 seconds) fails to
completely inactivate proteinases produced by some species and strains of Pseudomonas and other
psychrotrophs. UHT-treated milk can thus be spoiled by the residual activity of the proteinases
during storage. Lipases are only partially inactivated by pasteurization or by heating (in cream)
at 90°C for two minutes. They are generally inactivated by UHT treatment. Lipases from some
Pseudomonas strains retain sufficient activity even after heating at 100°C for 10 minutes.
Heat-stable proteinases of psychrotrophic bacteria differ in their substrate specificity and rate
of substrate degradation. Proteinases of Pseudomonas spp. preferentially degrade casein of milk
by a different mechanism. Proteinases from some species or strains initially degrade β-casein,
whereas proteinases from other species or strains initially degrade k-casein. With time, they
can also degrade other casein fractions. Proteinases from psychrotrophic species or strains of
Flavobacterium, Aeromonas, and Serratia also showed initial differences in the degradation of βand k-caseins. α-Casein is degraded last by all strains.
Lipases of psychrotrophic bacteria differ in their specificity toward lipids. A lipase from a strain
of Pse. fragi specifically hydrolyzes fatty acids from positions 1 and 3 of the triglycerides, whereas
another lipase from a second strain hydrolyzes only at position 1. Similar specificities have been
observed with extracellular lipases from Pse. fluorescens. Some lipases of psychrotrophic bacteria
equally hydrolyze all three fatty acids.
Among the phospholipases produced by psychrotrophic Gram-negative bacteria, phospholipase C from Pseudomonas spp. has relatively high heat stability. It is not destroyed by pasteurization.
Phospholipase C from Pse. fluorescens is more active against phosphatidylethanolamine than other
phospholipids.
Spoilage of Foods with Heat-Stable Microbial enzymes
The presence of heat-stable extracellular enzymes of psychrotrophic bacteria in raw milk can cause
spoilage of dairy products made from it. In addition, when these dairy products are used as ingredients to make other food products, the action of heat-stable enzymes can also reduce their acceptance
qualities. Several examples are used here to emphasize the spoilage potential of these enzymes. 8,9
Pasteurized Milk
Heat-stable proteinases and lipases of psychrotrophic bacteria are not inactivated by pasteurization
and can cause proteolysis of casein and lipolysis of milk lipids to produce flavor defects. However,
raw milk, meat, fish, and other food products. During refrigerated storage, psychrotrophs are able
to grow and produce heat-stable enzymes in foods. 6,7
Production of heat-stable proteinases and lipases in milk by some of these bacteria, especially
Pseudomonas spp., and the characteristics of these enzymes have been well studied. The results show
that different species and strains produce proteinases and lipases that differ in molecular weight and
activity. Thus, a highly active proteinase or lipase produced by a Pseudomonas strain can produce
extensive proteolysis or lipolysis, even at 10 5–6 cells/mL milk. But another strain may need to reach
to 10 9 cells/mL to produce similar changes. The enzymes can be produced in raw milk at refrigerated temperatures (1°C–7°C) in sufficient quantities to hydrolyze proteins and lipids in detectable
levels within 3–7 days. The catalytic activity of the proteinases is highest between pH 6.0 and 7.0
with a pH range between 5.0 and 9.0. Pasteurization of milk (at 63°C for 30 minutes or 71°C for
15 seconds) results in a loss of 6%–36% activity and, even after heating at 121°C for 10 minutes,
some activity of the proteinases is retained. UHT treatment (140°C–150°C for 1–5 seconds) fails to
completely inactivate proteinases produced by some species and strains of Pseudomonas and other
psychrotrophs. UHT-treated milk can thus be spoiled by the residual activity of the proteinases
during storage. Lipases are only partially inactivated by pasteurization or by heating (in cream)
at 90°C for two minutes. They are generally inactivated by UHT treatment. Lipases from some
Pseudomonas strains retain sufficient activity even after heating at 100°C for 10 minutes.
Heat-stable proteinases of psychrotrophic bacteria differ in their substrate specificity and rate
of substrate degradation. Proteinases of Pseudomonas spp. preferentially degrade casein of milk
by a different mechanism. Proteinases from some species or strains initially degrade β-casein,
whereas proteinases from other species or strains initially degrade k-casein. With time, they
can also degrade other casein fractions. Proteinases from psychrotrophic species or strains of
Flavobacterium, Aeromonas, and Serratia also showed initial differences in the degradation of βand k-caseins. α-Casein is degraded last by all strains.
Lipases of psychrotrophic bacteria differ in their specificity toward lipids. A lipase from a strain
of Pse. fragi specifically hydrolyzes fatty acids from positions 1 and 3 of the triglycerides, whereas
another lipase from a second strain hydrolyzes only at position 1. Similar specificities have been
observed with extracellular lipases from Pse. fluorescens. Some lipases of psychrotrophic bacteria
equally hydrolyze all three fatty acids.
Among the phospholipases produced by psychrotrophic Gram-negative bacteria, phospholipase C from Pseudomonas spp. has relatively high heat stability. It is not destroyed by pasteurization.
Phospholipase C from Pse. fluorescens is more active against phosphatidylethanolamine than other
phospholipids.
Spoilage of Foods with Heat-Stable Microbial enzymes
The presence of heat-stable extracellular enzymes of psychrotrophic bacteria in raw milk can cause
spoilage of dairy products made from it. In addition, when these dairy products are used as ingredients to make other food products, the action of heat-stable enzymes can also reduce their acceptance
qualities. Several examples are used here to emphasize the spoilage potential of these enzymes. 8,9
Pasteurized Milk
Heat-stable proteinases and lipases of psychrotrophic bacteria are not inactivated by pasteurization
and can cause proteolysis of casein and lipolysis of milk lipids to produce flavor defects. However,