Food Biopreservatives of Microbial Origin, Bacteriocin, and Nanotechnology ◾ 223
position 27 and histidine at position 31 with lysine residues. Sensitivity of microcin J25, produced
by Escherichia coli, to chymotrypsin was achieved by substitution of glycine at position 12 with
tyrosine without affecting its antimicrobial activity. Introduction of a disulphide bridge in the
C-terminal end of sakacin P broadened its antimicrobial spectrum. A chimeric bacteriocin consisting of the C-terminal half of pediocin and the N-terminal half of enterocin A yielded improved
antimicrobial activity against Leuconostoc lactis, a dairy product spoilage bacterium. Continued
efforts are needed in developing desirable bacteriocins through bioengineering to enhance antimicrobial spectrum and stability during food preservation.
Production and Purification
Bacteriocin production in lactic acid bacteria is directly related to the cell mass. Generally, the
parameters that help generate more cell mass produce more bacteriocin molecules. The parameters include nutritional composition, initial and terminal pH and O-R potential of a broth, and
incubation temperature and time. The species and the strains of a species growing under similar
conditions differ greatly in the amount of bacteriocin production. In general, production of nisin
A or pediocin AcH is much higher than leucocin or sakacin A. A nutritionally rich medium is
always better, and, for many bacteriocins, growing the strains in a fermentor under a controlled
terminal pH produces more bacteriocin. For example, nisin A production is much higher at pH
6.0, but pediocin AcH production is much higher at a lower terminal pH of 3.6. It is important to
determine the optimum parameters for growth and bacteriocin production for an unknown strain
to obtain a high yield of bacteriocin.
Bacteriocin molecules following secretion by a producer strain remain either adsorbed on the cell
surface or free in the medium, depending on the pH of the environment. More molecules remain
free in the environment at pH 1.5–2.0, whereas at pH 6.0–7.0, they remain bound to the cell surface.
Based on this pH-dependent adsorption-desorption of bacteriocin on the cell surface of a producer
strain, an effective and easy method has been developed to purify bacteriocin molecules in large
amounts. A second method of purification involves precipitation of the molecules by ammonium sulfate followed by stepwise gel filtration. This gives a highly purified preparation but in small amounts.
Applications 8–11,15,18
Bacteriocins of food-grade lactic acid bacteria are considered safe food biopreservatives and have
the potential to be used to kill sensitive Gram-positive food spoilage and food-borne pathogenic
bacteria. Among the bacteriocins, only nisin has received regulatory approval for use in select
foods. In 1969, the Joint FAO/WHO Expert Committee on Food Additives approved its use and
gave it the food additive number 234. The US FDA gave nisin GRAS status and reported nisin as
safe for human consumption at 2.94 mg/day and is approved for use in cooked meat and poultry
products at 5.5 mg nisin/kg (FDA GRAS Notice No. GRN 000065). In foods that can contain
injured Gram-negative bacteria, bacteriocins can also be effectively used to kill them. They are
more effective when used in minimally heat-processed foods in suitable combinations of two or
more (e.g., nisin and pediocin together). Table 17.5 presents the effectiveness of a preparation containing nisin and pediocin against pathogenic and spoilage bacteria in processed meat products
during refrigerated storage. The results show that the bacteriocin preparation effectively reduced
the Gram-positive spoilage (Leu. mesenteroides) and pathogenic (Lis. monocytogenes) bacteria as well
as Gram-negative pathogens (Salmonella and Esc. coli O15:H7) during six weeks of storage at 4°C.
Bacteriocins are examined in combination with other antimicrobials, such as acids, salts, spice
position 27 and histidine at position 31 with lysine residues. Sensitivity of microcin J25, produced
by Escherichia coli, to chymotrypsin was achieved by substitution of glycine at position 12 with
tyrosine without affecting its antimicrobial activity. Introduction of a disulphide bridge in the
C-terminal end of sakacin P broadened its antimicrobial spectrum. A chimeric bacteriocin consisting of the C-terminal half of pediocin and the N-terminal half of enterocin A yielded improved
antimicrobial activity against Leuconostoc lactis, a dairy product spoilage bacterium. Continued
efforts are needed in developing desirable bacteriocins through bioengineering to enhance antimicrobial spectrum and stability during food preservation.
Production and Purification
Bacteriocin production in lactic acid bacteria is directly related to the cell mass. Generally, the
parameters that help generate more cell mass produce more bacteriocin molecules. The parameters include nutritional composition, initial and terminal pH and O-R potential of a broth, and
incubation temperature and time. The species and the strains of a species growing under similar
conditions differ greatly in the amount of bacteriocin production. In general, production of nisin
A or pediocin AcH is much higher than leucocin or sakacin A. A nutritionally rich medium is
always better, and, for many bacteriocins, growing the strains in a fermentor under a controlled
terminal pH produces more bacteriocin. For example, nisin A production is much higher at pH
6.0, but pediocin AcH production is much higher at a lower terminal pH of 3.6. It is important to
determine the optimum parameters for growth and bacteriocin production for an unknown strain
to obtain a high yield of bacteriocin.
Bacteriocin molecules following secretion by a producer strain remain either adsorbed on the cell
surface or free in the medium, depending on the pH of the environment. More molecules remain
free in the environment at pH 1.5–2.0, whereas at pH 6.0–7.0, they remain bound to the cell surface.
Based on this pH-dependent adsorption-desorption of bacteriocin on the cell surface of a producer
strain, an effective and easy method has been developed to purify bacteriocin molecules in large
amounts. A second method of purification involves precipitation of the molecules by ammonium sulfate followed by stepwise gel filtration. This gives a highly purified preparation but in small amounts.
Applications 8–11,15,18
Bacteriocins of food-grade lactic acid bacteria are considered safe food biopreservatives and have
the potential to be used to kill sensitive Gram-positive food spoilage and food-borne pathogenic
bacteria. Among the bacteriocins, only nisin has received regulatory approval for use in select
foods. In 1969, the Joint FAO/WHO Expert Committee on Food Additives approved its use and
gave it the food additive number 234. The US FDA gave nisin GRAS status and reported nisin as
safe for human consumption at 2.94 mg/day and is approved for use in cooked meat and poultry
products at 5.5 mg nisin/kg (FDA GRAS Notice No. GRN 000065). In foods that can contain
injured Gram-negative bacteria, bacteriocins can also be effectively used to kill them. They are
more effective when used in minimally heat-processed foods in suitable combinations of two or
more (e.g., nisin and pediocin together). Table 17.5 presents the effectiveness of a preparation containing nisin and pediocin against pathogenic and spoilage bacteria in processed meat products
during refrigerated storage. The results show that the bacteriocin preparation effectively reduced
the Gram-positive spoilage (Leu. mesenteroides) and pathogenic (Lis. monocytogenes) bacteria as well
as Gram-negative pathogens (Salmonella and Esc. coli O15:H7) during six weeks of storage at 4°C.
Bacteriocins are examined in combination with other antimicrobials, such as acids, salts, spice