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This thesis investigates a newly developed and patented technology for its ability to inactivate spore- forming bacteria and non-spore-forming microorganisms. This new technology “Pressure Assisted Thermal Sterilization©” (PATS) is based on the theory of the thermal expansion of liquids. The efficiency of inactivating spore-forming and non-spore-forming microorganisms by PATS was compared with the thermal treatment alone. A combination treatment consisting of high pressure processing and gaseous carbon dioxide was also investigated for its ability to inactivate bacterial spores in model and real food matrices. The structural damage caused by treatments to the spores and non-spore-forming bacteria was assessed by scanning electron microscopy. Geobacillus stearothermophilus spores suspended in Milli-Q water, UHT milk and pumpkin soup, treated by PATS were found to have significantly lower decimal reduction times (D values) compared with the thermal treatment alone. Spores suspended in UHT milk were more heat resistant compared with those in Milli-Q water and pumpkin soup. Bacillus cereus spores suspended in Milli-Q water and pumpkin soup treated with PATS were more effectively inactivated compared with spores treated by the thermal treatment alone. Clostridium botulinum spores in saline buffer subjected to PATS treatment were inactivated more effectively compared with the thermal treatment alone. Overall, the results show that PATS was a better processing technique for inactivation of bacterial spores compared with thermal treatment alone. However, PATS had no added benefit in inactivating the non-spore-forming bacteria Escherichia coli and Saccharomyces cerevisiae cells compared with the thermal treatment. A shelf life study showed that B. cereus spores in pumpkin soup retained a low spore count (<5 LogCFU/mL) for approximately 40 days in 30oC storage after treatment with PATS. No additional degradation of colour pigments of pumpkin soup and model pumpkin juice was observed following PATS compared with the thermal treatment. Spore-forming microorganisms can be resistant to pressure treatment alone, which limits the application of high pressure processing (HPP). Therefore, a combination approach was investigated. The mechanism of inactivating spores by combining HPP with other treatments is that the pressure assists in spore germination. Then a secondary treatment (thermal or CO2 gas) can be used to inactivate the germinated spores. A combined application of HPP and a consecutive CO2 treatment was investigated for the efficiency of spore inactivation. Results showed that HPP (200 MPa for 30 min) followed by a CO2 treatment inactivated Bacillus subtilis 168 in nutrient broth, tomato juice and liquid whole egg by 2.5, 1.0 and 1.5 LogCFU/mL respectively. These results indicated that this technique is inadequate for practical use. Scanning electron micrographs showed that pressure processing of B. subtilis 168 and B. subtilis natto spores resulted in deformation of the spore structure. This structural deformation of spores may have been due to water absorption during HPP and subsequent release upon decompression. PATS treated G. stearothermophilus and B. cereus spores were more severely damaged compared with the same spores which underwent thermal treatment alone. However, the extent to which E. coli and S. cerevisiae cells were damaged by both PATS and thermal treatment was similar.

This thesis investigates a newly developed and patented technology for its ability to inactivate spore- forming bacteria and non-spore-forming microorganisms. This new technology “Pressure Assisted Thermal Sterilization©” (PATS) is based on the theory of the thermal expansion of liquids. The efficiency of inactivating spore-forming and non-spore-forming microorganisms by PATS was compared with the thermal treatment alone. A combination treatment consisting of high pressure processing and gaseous carbon dioxide was also investigated for its ability to inactivate bacterial spores in model and real food matrices. The structural damage caused by treatments to the spores and non-spore-forming bacteria was assessed by scanning electron microscopy. Geobacillus stearothermophilus spores suspended in Milli-Q water, UHT milk and pumpkin soup, treated by PATS were found to have significantly lower decimal reduction times (D values) compared with the thermal treatment alone. Spores suspended in UHT milk were more heat resistant compared with those in Milli-Q water and pumpkin soup. Bacillus cereus spores suspended in Milli-Q water and pumpkin soup treated with PATS were more effectively inactivated compared with spores treated by the thermal treatment alone. Clostridium botulinum spores in saline buffer subjected to PATS treatment were inactivated more effectively compared with the thermal treatment alone. Overall, the results show that PATS was a better processing technique for inactivation of bacterial spores compared with thermal treatment alone. However, PATS had no added benefit in inactivating the non-spore-forming bacteria Escherichia coli and Saccharomyces cerevisiae cells compared with the thermal treatment. A shelf life study showed that B. cereus spores in pumpkin soup retained a low spore count (<5 LogCFU/mL) for approximately 40 days in 30oC storage after treatment with PATS. No additional degradation of colour pigments of pumpkin soup and model pumpkin juice was observed following PATS compared with the thermal treatment. Spore-forming microorganisms can be resistant to pressure treatment alone, which limits the application of high pressure processing (HPP). Therefore, a combination approach was investigated. The mechanism of inactivating spores by combining HPP with other treatments is that the pressure assists in spore germination. Then a secondary treatment (thermal or CO2 gas) can be used to inactivate the germinated spores. A combined application of HPP and a consecutive CO2 treatment was investigated for the efficiency of spore inactivation. Results showed that HPP (200 MPa for 30 min) followed by a CO2 treatment inactivated Bacillus subtilis 168 in nutrient broth, tomato juice and liquid whole egg by 2.5, 1.0 and 1.5 LogCFU/mL respectively. These results indicated that this technique is inadequate for practical use. Scanning electron micrographs showed that pressure processing of B. subtilis 168 and B. subtilis natto spores resulted in deformation of the spore structure. This structural deformation of spores may have been due to water absorption during HPP and subsequent release upon decompression. PATS treated G. stearothermophilus and B. cereus spores were more severely damaged compared with the same spores which underwent thermal treatment alone. However, the extent to which E. coli and S. cerevisiae cells were damaged by both PATS and thermal treatment was similar.

This thesis investigates a newly developed and patented technology for its ability to inactivate spore- forming bacteria and non-spore-forming microorganisms. This new technology “Pressure Assisted Thermal Sterilization©” (PATS) is based on the theory of the thermal expansion of liquids. The efficiency of inactivating spore-forming and non-spore-forming microorganisms by PATS was compared with the thermal treatment alone. A combination treatment consisting of high pressure processing and gaseous carbon dioxide was also investigated for its ability to inactivate bacterial spores in model and real food matrices. The structural damage caused by treatments to the spores and non-spore-forming bacteria was assessed by scanning electron microscopy. Geobacillus stearothermophilus spores suspended in Milli-Q water, UHT milk and pumpkin soup, treated by PATS were found to have significantly lower decimal reduction times (D values) compared with the thermal treatment alone. Spores suspended in UHT milk were more heat resistant compared with those in Milli-Q water and pumpkin soup. Bacillus cereus spores suspended in Milli-Q water and pumpkin soup treated with PATS were more effectively inactivated compared with spores treated by the thermal treatment alone. Clostridium botulinum spores in saline buffer subjected to PATS treatment were inactivated more effectively compared with the thermal treatment alone. Overall, the results show that PATS was a better processing technique for inactivation of bacterial spores compared with thermal treatment alone. However, PATS had no added benefit in inactivating the non-spore-forming bacteria Escherichia coli and Saccharomyces cerevisiae cells compared with the thermal treatment. A shelf life study showed that B. cereus spores in pumpkin soup retained a low spore count (<5 LogCFU/mL) for approximately 40 days in 30oC storage after treatment with PATS. No additional degradation of colour pigments of pumpkin soup and model pumpkin juice was observed following PATS compared with the thermal treatment. Spore-forming microorganisms can be resistant to pressure treatment alone, which limits the application of high pressure processing (HPP). Therefore, a combination approach was investigated. The mechanism of inactivating spores by combining HPP with other treatments is that the pressure assists in spore germination. Then a secondary treatment (thermal or CO2 gas) can be used to inactivate the germinated spores. A combined application of HPP and a consecutive CO2 treatment was investigated for the efficiency of spore inactivation. Results showed that HPP (200 MPa for 30 min) followed by a CO2 treatment inactivated Bacillus subtilis 168 in nutrient broth, tomato juice and liquid whole egg by 2.5, 1.0 and 1.5 LogCFU/mL respectively. These results indicated that this technique is inadequate for practical use. Scanning electron micrographs showed that pressure processing of B. subtilis 168 and B. subtilis natto spores resulted in deformation of the spore structure. This structural deformation of spores may have been due to water absorption during HPP and subsequent release upon decompression. PATS treated G. stearothermophilus and B. cereus spores were more severely damaged compared with the same spores which underwent thermal treatment alone. However, the extent to which E. coli and S. cerevisiae cells were damaged by both PATS and thermal treatment was similar.

This thesis investigates a newly developed and patented technology for its ability to inactivate spore- forming bacteria and non-spore-forming microorganisms. This new technology “Pressure Assisted Thermal Sterilization©” (PATS) is based on the theory of the thermal expansion of liquids. The efficiency of inactivating spore-forming and non-spore-forming microorganisms by PATS was compared with the thermal treatment alone. A combination treatment consisting of high pressure processing and gaseous carbon dioxide was also investigated for its ability to inactivate bacterial spores in model and real food matrices. The structural damage caused by treatments to the spores and non-spore-forming bacteria was assessed by scanning electron microscopy. Geobacillus stearothermophilus spores suspended in Milli-Q water, UHT milk and pumpkin soup, treated by PATS were found to have significantly lower decimal reduction times (D values) compared with the thermal treatment alone. Spores suspended in UHT milk were more heat resistant compared with those in Milli-Q water and pumpkin soup. Bacillus cereus spores suspended in Milli-Q water and pumpkin soup treated with PATS were more effectively inactivated compared with spores treated by the thermal treatment alone. Clostridium botulinum spores in saline buffer subjected to PATS treatment were inactivated more effectively compared with the thermal treatment alone. Overall, the results show that PATS was a better processing technique for inactivation of bacterial spores compared with thermal treatment alone. However, PATS had no added benefit in inactivating the non-spore-forming bacteria Escherichia coli and Saccharomyces cerevisiae cells compared with the thermal treatment. A shelf life study showed that B. cereus spores in pumpkin soup retained a low spore count (<5 LogCFU/mL) for approximately 40 days in 30oC storage after treatment with PATS. No additional degradation of colour pigments of pumpkin soup and model pumpkin juice was observed following PATS compared with the thermal treatment. Spore-forming microorganisms can be resistant to pressure treatment alone, which limits the application of high pressure processing (HPP). Therefore, a combination approach was investigated. The mechanism of inactivating spores by combining HPP with other treatments is that the pressure assists in spore germination. Then a secondary treatment (thermal or CO2 gas) can be used to inactivate the germinated spores. A combined application of HPP and a consecutive CO2 treatment was investigated for the efficiency of spore inactivation. Results showed that HPP (200 MPa for 30 min) followed by a CO2 treatment inactivated Bacillus subtilis 168 in nutrient broth, tomato juice and liquid whole egg by 2.5, 1.0 and 1.5 LogCFU/mL respectively. These results indicated that this technique is inadequate for practical use. Scanning electron micrographs showed that pressure processing of B. subtilis 168 and B. subtilis natto spores resulted in deformation of the spore structure. This structural deformation of spores may have been due to water absorption during HPP and subsequent release upon decompression. PATS treated G. stearothermophilus and B. cereus spores were more severely damaged compared with the same spores which underwent thermal treatment alone. However, the extent to which E. coli and S. cerevisiae cells were damaged by both PATS and thermal treatment was similar.

O processamento em alta pressão é um método alternativo para a produção de materiais cerâmicos. Neste trabalho, pressões na ordem de 7,7 GPa e 2,5 GPa foram aplicadas em amostras, em diferentes temperaturas, que foram colocadas em uma célula de reação específica, gerando diferentes efeitos na formação de fases. A composição de fases foi analisada por difração de raios X e a evolução microestrutural, associada ao processamento em alta pressão, foi investigada por microscopia eletrônica por varredura em associação com a espectroscopia por dispersão de energia. Um analisador de resposta de frequência foi utilizado para obter as curvas ferroelétricas por espectroscopia de impedância eletroquímica. A utilização de alta pressão (2,5 GPa) possibilitou a obtenção de amostras de SrBi2Ta2O9 monofásicas com elevada densidade relativa, acima de 93%, após sinterização a uma temperatura de 900 °C. Essa temperatura é inferior às usualmente necessárias para obter alta densificação utilizando métodos convencionais de sinterização. Além disso, as amostras processadas em alta pressão apresentaram uma resposta dielétrica similar às amostras de SrBi2Ta2O9 sinterizadas por processos convencionais em temperaturas acima de 1000 ºC. / High-pressure processing is a very attractive approach for the production of ceramic materials. In this work, pressures about 7.7 GPa and 2.5 GPa were applied in SrBi2Ta2O9 samples at different temperatures placed in a specific reaction cell. X-ray diffraction was used to identify the different phases produced as a function of the processing conditions. The microstructural evolution, associated to the high-pressure processing, was investigated by scanning electron microscopy in association with energy dispersive spectroscopy. Frequency response analysis was used to obtain the ferroelectric curves by electrochemical impedance spectroscopy. A highly densified (> 93% of theoretical density) single-phase (SrBi2Ta2O9) sample was obtained after processing at 2.5 GPa and 900 ºC. This temperature is lower than those necessary to obtain high densification, when conventional sintering processes are employed. In addition, the samples produced by high pressure processing showed a dielectric response similar to SrBi2Ta2O9 samples sintered by conventional processes at temperatures above 1000 ºC.

Les endospores bactériennes sont l’une des formes les plus résistantes du vivant. Leur capacité à survivre aux traitements de décontamination et leur potentielle pathogénicité pose de réels problèmes aux industriels de l’agroalimentaire. L’usage de conservateurs ou de traitements thermiques reste aujourd’hui la seule solution pour empêcher leur développement dans les aliments. Cependant, les impacts négatifs de ces deux procédés sur les qualités nutritionnelles et la santé du consommateur poussent les industriels à se tourner vers des méthodes de décontamination alternatives.Le procédé de traitement par hautes pressions hydrostatiques (HP) est l’un des nouveaux procédés de décontamination athermique le plus rependu pour la pasteurisation des aliments. Cependant, ce procédé n’a que très peu d’effet sur les endospores et doit donc être combiné avec d’autres méthodes de décontamination. L’objectif de cette thèse a été d’évaluer le potentiel de la combinaison du traitement HP avec la biopréservation pour inactiver des spores thermorésistantes et pathogènes. Cette méthode de décontamination douce met en œuvre des bactéries lactiques protectrices ou les molécules antimicrobiennes qu’elles produisent, telles que la nisine. Des méthodes d’investigations globales telles que la microspectroscopie, la spectroscopie infrarouge et le dichroïsme circulaire, il a été démontré que les HP avaient un effet non négligeable sur l’effet antimicrobien de la nisine. En affectant les structures secondaires de cette protéine, la pression induit une diminution non négligeable de son action antimicrobienne. Cependant, présente dans le milieu de recouvrement de spores traitées en pression, la nisine peut induire une inactivation conséquente (>7 log) d’une population de spores thermophiles et pathogènes. Cette sensibilisation des spores à a nisine par les HP est due l’initiation des toutes premières phases de la germination des spores induites par la pression, non détectées par des méthodes d’analyses classiques. Ces résultats apportent ainsi de nouvelles données pour la compréhension de l’effet des HP sur les spores. Ils permettent aussi d’envisager l’utilisation conjointe des HP et de la nisine à l’échelle industrielle. / Bacterial endospores are one of the most resistant life form on earth. Their capacity to survive to decontamination processes and their potential pathogenicity represent a real problem for the food industry. Currently, the only way to prevent their development in foods is the application of thermal treatments or the use of preservatives. However, these two methods have negative impacts on the nutritional properties of foods and on the consumers’ health. High hydrostatic pressure (HP) is a non-thermal process widely used for commercial pasteurization of foods. However, this process has a very low effect on spores and has therefore to be combined with other decontamination processes to enhance its effectiveness. The objective of this work was to evaluate the potential of the use of biopreservation as an additional hurdle for the inactivation of thermoresistant and pathogenic foodborne spores by HP. Biopreservation is a gentle decontamination process involving protective culture or the antimicrobial agents they produce, like nisin. Thanks to global investigation methods such as microspectroscopy, infrared spectroscopy or circular dichroism, this study showed that HP treatment could affect the antimicrobial properties of nisin. By affecting the secondary structures of this protein, HP can induce a drastic drop in its antimicrobial activity. However when added in the recovery medium of HP-treated spores, nisin can induce their synergistic inactivation (> 7log). This HP-sensitization of spore to nisin is due to the induction of the very first steps of the germination process, usually not detected by the current methods of germination analysis. These results bring knew knowledges about the underlying mechanisms of spores germination under HP and gives new perspectives for the combined used of HP and nisin at the industrial scale.

This thesis discussed the non-thermal food processing technologies being used within the United States of America. The technologies discussed in this thesis are High- Pressure Processing (HHP), Pulsed Electric Field, Pulsed Light, Irradiation, Ultra Sound, Oscillating Magnetic Fields, and Cold Atmospheric Plasma. A survey was designed and conducted to study the major reasons behind a preference for a particular technology by the organization, and the limitations for not implementing specific technologies. The survey participants were management level, food scientists and, food technologists employed by food processing companies. The questionnaire consisted of ten questions related to demographics, current technology, barriers from other technologies, and research and development of new technologies. There were a total 223 respondents from various regions of the United States. The respondents had a wide array of industry experience. Of the respondents, 91% of the respondents had either a Bachelor’s Degree, Master’s Degree or Ph D. Thirty-six percent of the participants chose high pressure processing and 20 % chose pulsed electric as the most commonly used non-thermal food processing technologies. Rapidly increasing technologies included cold atmospheric plasma and oscillating magnetic fields. Seventyone percent mentioned the main driver for them to choose non-thermal food processing was better nutrient and sensory properties. As per the results, 41% of respondents believed the major limitations in implementing non-thermal food processing technologies was high investment. The results indicated the main drivers for innovation were equipment manufacturers and research. These researches were either academic or government funded.

Outgrowth of spoilage yeasts and moulds and post-processing acidification can limit the shelf-life of some fermented dairy products including fresh lactic curd cheeses. The possibility of using high pressure processing (HPP) for controlling these problems was investigated in a commercially manufactured fresh lactic curd cheese (pH 4.3-4.4) and fermented milk models (pH 4.3-6.5). The effects of HPP at 300 and 600 MPa on inactivation of glycolytic enzymes of lactic acid bacteria were also evaluated. Fresh cheeses made from pasteurised bovine milk using a commercial Lactococcus starter preparation were treated with high pressures ranging from 200 to 600 MPa (less than or equal to 22°C, 5 min) under vacuum packaging conditions and subsequently stored at 4°C for 8 weeks. Treatment at greater than or equal to 300 MPa substantially reduced the viable count of Lactococcus and effectively prevented the outgrowth of yeasts and moulds for 6 to 8 weeks without adversely affecting the sensory and textural attributes of the product. However, it had no significant effects (p less than 0.01) on variation of titratable acidity during storage. Fermented milk models were prepared by individually growing Lactococcus lactis subsp. lactis C10, Lactococcus lactis subsp. cremoris BK5, Streptococcus thermophilus TS1, Lactobacillus acidophilus 2400 and Lactobacillus delbrueckii subsp. bulgaricus 2517 in UHT skim milk and diluting the resulting fermented milk with UHT skim milk up to pH 6.5. Pressure treatment of the milk models at pH 5.2 resulted in substantial inhibition of post-processing acidification during storage and markedly reduced the viable count of Lactococcus at both 300 and 600 MPa and other bacteria only at 600 MPa. Treatment of the milk model at 600 MPa decreased the viable counts of Candida zeylanoides and Candida lipolytica (wildtype spoilage yeasts of lactic curd cheese, added as challenge cultures) from 105 CFU mL-1 to below the detection limit (log 0 CFU mL-1) at all pH levels tested (pH 4.3-6.5) and effectively controlled their outgrowth for 8 weeks. Treatment of milk model at 300 MPa had a similar effect only on C. zeylanoides. The viable count of C. lipolytica was reduced by 2.6, 2.4 and 2.3 logs by treatment at 300 MPa at pH levels of 4.3, 5.2 and 6.5, respectively, which subsequently recovered by 2.9, 2.8 and 3.2 logs within 3 weeks. Glycolytic enzymes of various starter bacteria showed different responses to pressure treatment. The lactate dehydrogenase in L. lactis subsp. lactis and Lb. acidophilus was quite resistant to pressures up to 600 MPa, but it was almost completely inactivated in S. thermophilus at pressure levels as low as 300 MPa. The â-galactosidase in Lb. acidophilus was more pressure stable than â-galactosidase in S. thermophilus and Phospho-â-galactosidase in L. lactis subsp. lactis. The findings of this study suggests HPP at 300-600 MPa as an effective method for controlling the outgrowth of some spoilage yeasts and moulds in fresh lactic curd cheeses. The results obtained with selected lactic acid bacteria in fermented milk models can be used to assist in establishing HPP operating parameters for development of new generation cultured dairy products, of reduced acidity and extended shelf-life.

O processamento em alta pressão é um método alternativo para a produção de materiais cerâmicos. Neste trabalho, pressões na ordem de 7,7 GPa e 2,5 GPa foram aplicadas em amostras, em diferentes temperaturas, que foram colocadas em uma célula de reação específica, gerando diferentes efeitos na formação de fases. A composição de fases foi analisada por difração de raios X e a evolução microestrutural, associada ao processamento em alta pressão, foi investigada por microscopia eletrônica por varredura em associação com a espectroscopia por dispersão de energia. Um analisador de resposta de frequência foi utilizado para obter as curvas ferroelétricas por espectroscopia de impedância eletroquímica. A utilização de alta pressão (2,5 GPa) possibilitou a obtenção de amostras de SrBi2Ta2O9 monofásicas com elevada densidade relativa, acima de 93%, após sinterização a uma temperatura de 900 °C. Essa temperatura é inferior às usualmente necessárias para obter alta densificação utilizando métodos convencionais de sinterização. Além disso, as amostras processadas em alta pressão apresentaram uma resposta dielétrica similar às amostras de SrBi2Ta2O9 sinterizadas por processos convencionais em temperaturas acima de 1000 ºC. / High-pressure processing is a very attractive approach for the production of ceramic materials. In this work, pressures about 7.7 GPa and 2.5 GPa were applied in SrBi2Ta2O9 samples at different temperatures placed in a specific reaction cell. X-ray diffraction was used to identify the different phases produced as a function of the processing conditions. The microstructural evolution, associated to the high-pressure processing, was investigated by scanning electron microscopy in association with energy dispersive spectroscopy. Frequency response analysis was used to obtain the ferroelectric curves by electrochemical impedance spectroscopy. A highly densified (> 93% of theoretical density) single-phase (SrBi2Ta2O9) sample was obtained after processing at 2.5 GPa and 900 ºC. This temperature is lower than those necessary to obtain high densification, when conventional sintering processes are employed. In addition, the samples produced by high pressure processing showed a dielectric response similar to SrBi2Ta2O9 samples sintered by conventional processes at temperatures above 1000 ºC.