The effect of using citric or acetic acid on survival of Listeria monocytogenes during fish protein recovery by isoelectric solubilization and precipitation process.

Isoelectric solubilization and precipitation (ISP) is a protein recovery process effective at reducing Listeria innocua, a nonpathogenic bacterium typically used as a surrogate for L. monocytogenes in recovered trout protein. The response of L. monocytogenes to ISP processing was determined and compared to the response of L. innocua. Headed and gutted rainbow trout were inoculated with L. monocytogenes (10.16 log CFU/g), homogenized, and pH-adjusted with granular citric acid (pH 2.0 and 2.5) or glacial acetic acid (pH 3.0 and 3.5). Proteins were solubilized and centrifugation was used to remove insoluble components (skin, insoluble protein, so on). The supernatant was returned to the protein isoelectric point (pH 5.5) with NaOH and centrifuged to remove precipitated protein. Microbial load was enumerated on both growth and selective media; recovery was not significantly different (P > 0.05). Surviving cells from each component (protein, insoluble, and water) were compared to initial inoculum numbers. Significant reductions were detected at all pH (P < 0.05). The greatest reductions were at pH 3.0 with acetic acid, with a mean log reduction of 3.03 in the combined components, and a 3.53 log reduction in the protein portion. Data were compared to results from a previous study using L. innocua. Significant differences (P < 0.05) in recovery were found between the 2 species at pH 2.0 and 3.0 with greater recovery of L. monocytogenes, regardless of processing pH or acid type. These results demonstrate the variability in resistance between species and indicate that L. innocua is not an appropriate surrogate for L. monocytogenes for ISP processing with organic acids.

Survival of Listeria innocua in rainbow trout protein recovered by isoelectric solubilization and precipitation with acetic and citric acids.

During mechanical fish processing, a substantial amount of protein is discarded as by-products. Isoelectric solubilization and precipitation (ISP) is a process that uses extreme pH shifts to solubilize and precipitate protein from by-products to recover previously discarded protein. Typically, strong acids are used for pH reduction, but these acids do not have a pasteurization effect (6 log reduction) on bacterial load; therefore, organic acids were used during ISP processing to test the impact on Listeria innocua concentrations. Headed and gutted rainbow trout (Oncorhynchus mykiss) were inoculated with L. innocua, homogenized, and brought to the target pH with granular citric acid (pH 2.0 and 2.5) or glacial acetic acid (pH 3.0 and 3.5). Proteins were solubilized for 10 min at 4°C, and insoluble components (e.g., skin and insoluble protein) were removed by centrifugation. The remaining solution was pH shifted to the protein isoelectric point (pH 5.5) with sodium hydroxide, and precipitated protein was separated from the water. Microbial cells for each component (proteins, insolubles, and water) were enumerated on modified Oxford agar (MOX) and tryptic soy agar with 6% yeast extract (TSAYE). The sums of the surviving cells from each component were compared with the initial inoculum levels. No significant differences were observed between results obtained from TSAYE and from MOX (P > 0.05). Significant reductions in microbial populations were detected, regardless of pH or acid type (P < 0.05). The greatest reduction was at pH 3.0 with glacial acetic acid, resulting in a mean reduction of 6.41 log CFU/g in the recovered protein and 5.88 log CFU/g in the combined components. These results demonstrate the antimicrobial potential of organic acids in ISP processing.

Use of electron beam radiation for the reduction of Salmonella enterica serovars typhimurium and Tennessee in peanut butter.

Peanut butter and peanut paste products were implicated as the vehicle of contamination in an outbreak of Salmonella Typhimurium, which began in September 2008, and in the November 2006 outbreak of Salmonella Tennessee. Therefore, this study evaluated the effectiveness of electron beam (e-beam) radiation for the reduction of Salmonella serovars Tennessee (ATCC 10722) and Typhimurium (ATCC 14028) in creamy peanut butter. Each strain was studied independently. Peanut butter samples were inoculated with approximately 8.0 log CFU/g of Salmonella, and exposed to e-beam doses ranging from 0 to 3.1 kGy. Doses were confirmed with film dosimetry. Survivors were enumerated by standard spread plating on nonselective tryptic soy agar (TSA) and selective xylose-lysine-desoxycholate agar (XLD) media. Salmonella Tennessee was more susceptible to e-beam radiation, with 5.00- and 6.75-log reduction of cells on TSA and XLD, respectively, at the approximate e-beam dose of 3.0 kGy. Salmonella Typhimurium was reduced by 4.19 and 4.85 log on TSA and XLD, respectively, at the approximate e-beam dose of 3.0 kGy. D(10)-values show that Salmonella Typhimurium was more resistant (0.82 +/- 0.02 and 0.73 +/- 0.01 kGy on TSA and XLD, respectively) than was Salmonella Tennessee (0.72 +/- 0.02 and 0.60 +/- 0.01 kGy on TSA and XLD, respectively) to e-beam radiation (P < 0.05). The recovery on growth and selective media were different (P < 0.05), indicating cell injury. The results of this study demonstrate that e-beam radiation may be an effective processing step for the nonthermal inactivation of Salmonella in peanut butter.

Survival of Escherichia coli after isoelectric solubilization and precipitation of fish protein.

Protein recovery for fish processing by-products utilizes extreme pH shifts for isoelectric solubilization and precipitation. The purpose of this study was to determine if Escherichia coli would survive exposure to the extreme pH shifts during the protein recovery process. Fresh rainbow trout were beheaded, gutted, and minced and then inoculated with approximately 10(9) CFU of E. coli ATCC 25922 per g, homogenized, and brought to the target pH of 2.0, 3.0, 11.5, or 12.5 by the addition of concentrated hydrochloric acid or sodium hydroxide to solubilize muscle proteins. The homogenate was blended and centrifuged to separate the lipid and insoluble components (bones, skin, insoluble protein, etc.) from the protein solution. The protein solution was subjected to a second pH shift (pH 5.5) resulting in protein precipitation that was recovered with centrifugation. Microbial analysis was conducted on each fraction (i.e., lipid, insoluble components, protein, and water) with selective and nonselective media. The sums of the surviving E. coli in these fractions were compared with the initial inoculum. The greatest total microbial reduction occurred when the pH was shifted to 12.5 (P < 0.05), i.e., a 4.4-log reduction of cells on nonselective media and a 6.0-log reduction of cells on selective media. The use of selective and nonselective media showed that there was significant (P < 0.05) injury sustained by cells exposed to alkaline treatment (pH 11.5 and 12.5) in all fractions except the insoluble fraction at pH 11.5. Increasing the exposure time or the pH may result in greater bacterial reductions in the recovered protein.

Survival of Listeria innocua after isoelectric solubilization and precipitation of fish protein.

Protein wasted by the disposal of fish processing by-products may be recovered using isoelectric solubilization and precipitation. Extreme pH shifts are used to solubilize the protein and then it is recovered by precipitation and centrifugation. Microbial survival after this process is unknown; therefore, the purpose was to see if Listeria innocua would survive extreme pH shifts during the protein recovery process. Fresh rainbow trout fillets were inoculated with L. innocua, homogenized, and brought to the target pH of 2, 3, 11.5, or 12.5 by the addition of concentrated hydrochloric acid or sodium hydroxide. The proteins were allowed to solubilize at 4 degrees C for 10 min, centrifuged, and the lipid and insoluble components (bones, skin, insoluble protein, and so on) were removed. A 2nd pH shift (pH 5.5) and centrifugation was used to separate the precipitating protein and water fractions. Each constituent (lipid, protein, water, insoluble components) was analyzed for bacterial content using growth and selective media. The sums of the surviving L. innocua in these constituents were compared to the initial inoculum. There were no significant differences in recovery on growth or selective media (P > 0.05). The greatest loss occurred when the pH was shifted to 2, with a 3.1-log reduction in the combined fractions of the trout fillets and a 3.8-log reduction in the protein fraction. There were no significant losses when the pH was adjusted to 11.5 (P > 0.05). Future studies will continue to look at the effects of using organic acid, rather than inorganic, for protein solubilization.

Amino acid and mineral composition of protein and other components and their recovery yields from whole Antarctic krill (Euphausia superba) using isoelectric solubilization/precipitation.

Proteins and insolubles were recovered from whole Antarctic krill via novel isoelectric solubilization/precipitation using different pH treatments. The protein recovery yield was 45% to 50% (dry basis). The recovered proteins had higher (P < 0.05) content of essential amino acids (EAAs) and non-EAAs as well as higher (P < 0.05) ratio of total EAA/total AA than whole krill. The EAAs constituted almost 50% of total AAs. The least extreme pH treatments (pHs 3 and 12) yielded highest (P < 0.05) content of EAAs. The quality of recovered proteins was high based on EAAs meeting FAO/WHO/UNU recommendations for adults and infants. The basic pH yielded proteins with the lowest (P < 0.05) amount of minerals and the highest (P < 0.05) amount of Ca, P, and Mg in the insolubles when compared to the acidic treatments. However, both basic and acidic treatments effectively removed minerals from recovered proteins without the removal of the exoskeleton before processing. Therefore, besides high-quality proteins, the insolubles may provide a mineral supplement in the animal diet.

Inactivation kinetics of Escherichia coli by pulsed electron beam.

A novel and compact low-energy (keV) high-power pulsed electron beam (e-beam) that utilizes a secondary emission electron gun (SEEG) was designed and constructed. Escherichia coli JM 109 at a concentration of 10(6) CFU/mL was spread-plated on Luria-Bertani (LB) medium and subjected to the SEEG e-beam. The e-beam was administered as 1 or 5 pulses. The duration of a single pulse was constant at 5 micros, e-beam current density was constant at 25 mA/cm2, and e-beam energy varied between 60 and 82.5 keV. Following treatment with the SEEG e-beam, survivors of the irradiated E. coli samples were enumerated by a standard 10-fold dilution and spread-plated. The survivor curves were plotted on logarithmic scale as a function of e-beam dose. The D10-values were calculated as a negative reciprocal of the slope of the survivor curves. The D10-values for E. coli inactivated with 1- and 5-pulse SEEG e-beam were 0.0026 and 0.0217 Gy, respectively. These D10-values were considerably lower than published D10-values for E. coli inactivated with conventional high-energy continuous e-beam, likely due to shorter exposure time (t), greater current density (J), and a pulse mode of the SEEG e-beam. The SEEG e-beam showed promising results for microbial inactivation in a nonthermal manner; however, due to low energy of the SEEG e-beam, current applications are limited to surface decontamination. The SEEG e-beam may be an efficient processing step for surface inactivation of food-borne pathogens on ready-to-eat products, including fresh and leafy vegetables.

Amino acid, fatty acid, and mineral profiles of materials recovered from rainbow trout (Oncorhynchus mykiss) processing by-products using isoelectric solubilization/precipitation.

Protein, lipid, and insolubles (bones, skin, scales, fins, insoluble protein, and more) were recovered from rainbow trout processing by-products by means of isoelectric solubilization/precipitation at basic pH and acidic pH. Isoelectric solubilization/precipitation of the trout processing by-products resulted in the recovery of protein that was higher (P < 0.05) in essential amino acids (EAAs), non-EAAs, and total EAA/total AA ratio when compared to the processing by-products. Basic pH treatments yielded a higher (P < 0.05) content of EAAs than the acidic pH treatments. Nutritional quality of the recovered protein was high based on EAAs meeting the FAO/WHO/UNU recommendations for adults. The presence of omega-3 and omega-6 fatty acids (omega-3, omega-6 FAs) and the omega-3/omega-6 ratio in the recovered lipids were similar to the trout processing by-products, indicating that the pH treatments had no effect on these FAs. Ca and P contents of the processing by-products exceeded the recommended dietary allowances (RDA), but Fe and Mg did not. Basic pH treatments yielded protein with the lowest (P < 0.05) amount of minerals and the highest (P < 0.05) amount of Ca, P, and Mg in the insolubles when compared to acidic pH. The isoelectric solubilization/precipitation of the processing by-products effectively removed minerals from the recovered protein without removal of the bones, skin, scales, fins, and so on, prior to processing. The results indicated that isoelectric solubilization/precipitation, particularly at basic pH, permitted recovery of high-quality protein and lipids from fish processing by-products for human food uses; also, the recovered insolubles may be used in animal feeds as a source of minerals.