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Thesis (MScFoodSc)--Stellenbosch University, 2014. / ENGLISH ABSTRACT: Fresh ready-to-cook fish fillets are prone to rapid loss of freshness and other quality attributes, as well as accelerated growth of spoilage micro-organisms under sub-optimal storage conditions. Cape hake (Merluccius capensis) is an important seafood in South Africa; however, rapid loss of quality and eventual spoilage is a problem limiting the economic potential. Thus, the aim of this study was to investigate the effects of active (40% CO2 + 30% O2 + 30% N2) and passive (0.039% CO2 + 20.95% O2 + 78% N2) modified atmosphere packaging (MAP) under different storage temperatures (0°C, 4°C, and 8°C) on the quality attributes of Cape hake (Merluccius capensis) fish fillets. This was achieved by investigating the effects of MAP (with or without absorbent pads) and storage temperature on quality attributes (microbial, physicochemical and sensory), changes in composition and concentration of volatile compounds (VOCs) and shelf life of Cape hake fillets. Modified atmosphere packaging, storage temperature and the use of absorbent pads had a significant (p < 0.05) impacts on physicochemical properties of Cape hake fillets during refrigerated storage. Highest storage temperature (8°C) led to accelerated deterioration of packaged Cape hake fillets. Generally, active MAP better maintained the quality attributes of Cape hake than passive MAP at 0°C and 4°C. Headspace gas composition of O2 and CO2 were significantly influenced by the storage time, temperature, MAP conditions and their interactions (p < 0.05). Irrespective of storage temperature, active-MA packaged fillets had lower pH values in comparison to fillets stored under passive-MAP. Drip loss was higher in active-MA fillets packaged without absorbent pad. Passive-MAP fillets did not show any drip loss. Absorbent pad was used to add value to MAP storage as MAP resulted in drip. The use of absorbent pad combined with low storage temperature maintained the firmness of hake fillets, across all temperatures. The interaction of MAP, absorbent pad and storage temperature had a significant effect on the aerobic mesophillic bacteria counts. Based on the aerobic mesophillic bacteria count fillets stored under active-MAP at 0°C (5.2 log cfu/g) was limited to day 12, while the fillet stored under passive-MAP at 0°C (log cfu/g) was limited to greater than day 3. Overall sensory acceptability of fillets decreased with increase in storage temperature across all treatments. Additionally, MAP had a significant (p < 0.05) impact on sensory attributes such as appearance and odour acceptability, with active-MA packaged fillets stored at 0°C having highest overall acceptability. A total of 16 volatiles were identified in Cape hake fillets, including 4 primary VOCs and 12 secondary VOCs. The VOCs associated with spoilage include tri-methylamine (TMA) (ammonia like), esters (sickeningly sweet) and sulphur group (putrid). MAP had a significant (p< 0.05) influence on volatile composition and concentration. Active-MA packaged fillets performed better during storage and had lower TMA value of 0.85% on day 12 in comparison with 7.22% under passive-MAP on day 6 at 0°C. The results obtained demonstrated that changes in volatile compounds were significantly (p< 0.05) influenced by storage duration, temperature and MAP. The development of high levels of VOCs and off-odour corresponded with high aerobic mesophillic bacteria count (≥ 5.5 log cfu/g). Based on these developments the storage life of Cape hake fillets packaged under active-MAP with absorbent pad and stored at 0°C was limited to 12 d, while the passive-MAP (control) fillets stored at 0°C was limited to 3 d. The use of active-MAP, in combination with absorbent pads and 0°C storage in addition to good hygienic practices, was effective in maintaining the postharvest quality of Cape hake fish fillets and led to higher shelf life. / AFRIKAANSE OPSOMMING: Vars, gereed-vir-kook vis filette is geneig om gou hulle varsheid en ander gehalte kenmerke te verloor en die vervoegde groei van mikro-organismes wat bederf tydens stoor veroorsaak, vind plaas. Kaapse stokvis (Merluccius capensis) is in Suid-Afrika ‘n gewilde seekos maar die ekonomiese potensiaal daarvan word deur die feit dat dit so gou in gehalte afneem en bederf, beperk. Dus word daar met hierdie studie gepoog om die effek van aktiewe (40% CO2 + 30% O2 + 30% N2) en passiewe (0.039% CO2 + 20.95% O2 + 78% N2) aangepasde verpakking (MAP) onder verskillende stoortemperature (0°C, 4°C, en 8°C) op die gehalte kenmerke van Kaapse stokvis (Merluccius capensis) filette te ondersoek. Dit is gedoen deur om die effek van MAP (met of sonder kussinkies) en stoortemperatuur op die gehalte kenmerke (mikrobies, fisiochemies en sensories) asook veranderinge in komposisie en konsentrasie van vlugtige samestellings (VOCs) en die raklewe van Kaapse stokvis filette te ondersoek. Aangepasde atmosfeer verpakking, stoortemperatuur en die gebruik van absorberende kussinkies het ‘n groot impak (p < 0.05) op die fisiochemiese kenmerke van Kaapse stokvis tydens stoor in yskaste gehad. Hoë stoortemperature (8°C) het aanleiding gegee tot die vinnige bederf van verpakte Kaapse stokvis filette. Oor die algemeen het aktiewe MAP die gehalte van die Kaapse stokvis filette teen 0°C and 4°C beter bewaar. Die komposisie van O2 en CO2 is heelwat deur stoortyd, temperatuur, MA toestande en die interaksies tussen bogenoemde, beïnvloed (p< 0.05). By alle temperature het aktiewe MA verpakte filette laer pH waardes getoon in vergeleke met filette wat in onder passiewe MA verpak is. Die drupverlies was hoër in aktiewe MA filette verpak sonder absorberende kussinkies. Passiewe MAP filette het nie enige drupverlies getoon nie. Absorberende kussinkies is gebruik om waarde by te voeg tot MAP stoor aaangesien MAP gelei het tot drup. By alle temperature het die gebruik van absorberende kussinkies tesame met lae stoortemperature bygedra tot die behoud van fermheid. Die interaksie van MAP, absorberende kussinkies, en stoortemperatuur het ‘n groot effek gehad op die aerobiese mesofiliese bakterië telling. Weens die aerobiese mesofiliese bakterië telling is stoor van filette onder aktiewe MAP teen 0°C (5.2 log cfu/g) beperk tot dag 12, terwyl filette gestoor onder passiewe MAP teen 0°C ( log cfu/g) beperk is tot dag 3. Oor die algemeen het die sensoriese aanneemlikheid van filette sonder inasgneming van die behandeling, verklein met ‘n toename in stoortemperature. MAP het ook ‘n groot impak op die sensoriese kenmerke soos voorkoms, reuk, en aktiewe MA verpakte fillets gestoor teen 0°C is oor die algemeen die aanneemlikste. ‘n Totaal van 16 vlugtige substanse is in Kaapse stokvis identifiseer. Dit het vier primêre VOCs en 12 sekondêre VOCs ingesluit. Die VOCs wat met bederf assosieer word, sluit tri-metilamien (TMA) (soos ammoniak), esters (soet) en die swael groep (smetterig) in. MAP het ‘n groot (p < 0.05) invloed op die vlugtige komposisie en konsentrasie. Aktiewe MA verpakte filette het beter tydens stoor presteer en het ‘n laer TMA waarde van 0.85% op dag 12 gehad, in vergelyking met 7.22 % onder passiewe MAP op dag 6 teen 0°C. Die resultate toon dat veranderinge in vlugtige samestellings grootliks beïnvloed word (p < 0.05) deur stoortyd, temperature en MAP. Die ontwikkeling van hoë vlakke van VOCs, slegte reuke en verlies aan varsheid gaan tesame met hoë aerobiese mesofiliese bakterië telling (≥ 5.5 log cfu/g). Gegrond op hierdie tellings is die stoorleeftyd van Kaapse stokvis filette beperk tot dag 12, terwyl passiewe MAP (die kontrole) filette gestoor teen 0°C, beperk is tot dag 3. Om op te som, die gebruik van aktiewe MAP tesame met absorberende kussinkies en 0°C stoor asook goeie higiëniese praktyk, kon die na-oes gehalte van Kaapse stokvis filette behou en het gelei tot ‘n langer rakleeftyd.

The current trend of increasing demand for minimally processed food requires more effective preservation technologies than are presently used. In this study, an investigation has been made into a novel strategy to control some common foodborne pathogens, and therefore, to provide an alternative means for enhancing the safety and extending the shelf lives of food products. Modified atmosphere is able to extend the shelf life of seafood and meat products. In this study, a simulated controlled atmosphere (CA) broth system was used to investigate the potential of a modified atmosphere rich in CO2 at a concentration of 40%, supplemented with N2, to control common foodborne pathogens, such as Listeria monocytogenes, Escherichia coli O157:H7, Salmonella enterica serovar Typhimurium, Staphylococcus aureus and Vibrio parahaemolyticus. Controlled atmosphere significantly reduced the exponential growth rates of all tested pathogens, while the effects on other growth parameters (eg. lag phase duration and maximum population density) depended on the individual species and the specific growth conditions. The CA significantly extended the lag phase durations of S. aureus and V. parahaemolyticus at 20 degrees C at both pH 6.3 and 6.8, and that of L. monocytogenes at both 7 degrees C and 20 degrees C, and at both pH 6.3 and 6.8. The CA also significantly lowered the maximum population densities of S. aureus and V. parahaemolyticus at 20 degrees C, at pH 6.3 and 6.8, S. Typhimurium at pH 6.8, and L. monocytogenes at pH 6.3 and 7 degrees C. E. coli O157:H7 and S. Typhimurium were more resistant to the inhibitory effect of the CA, while S. aureus and V. parahaemolyticus were most sensitive. The inhibitory effect of CA was due mainly to the extensions of the lag phase duration and the reduction of the exponential growth rates of the test pathogens. This study confirms other studies that CA as a means for food preservation provides potential to control foodborne pathogens and therefore enhance the safety of a food product. The use of lactic acid bacteria (LAB) in controlling spoilage microorganisms and pathogens in foods has been a popular research theme worldwide. In this study, the antimicrobial effects of 18 lactic acid bacteria strains were evaluated in vitro, with emphasis on the most effective strain, the newly characterised Lactobacillus reuteri DPC16. The results demonstrated antagonistic effects of many strains against L. monocytogenes, E. coli O157:H7, S. Typhimurium and S. aureus. L. reuteri DPC16 showed the strongest antimicrobial activity against the tested pathogens including both Gram-positive and Gram-negative bacteria. Co-cultivation of L. reuteri DPC16, and co-incubation of its spent culture supernatant (DPC16-SCS), with the pathogens have demonstrated that the antimicrobial effect is bactericidal and valid at pH 4 - 6.5 and at a temperature as low as 10 degrees C. Further characterisation of the antimicrobial effect of L. reuteri DPC16 showed it to be mainly due to the presence of reuterin (ß-hydroxypropionaldehyde), although lactic acid may have also played a role. These characteristics of L. reuteri DPC16 and its metabolite reuterin make it an unique and potent candidate as a biopreservative to control both Gram-positive and Gram-negative bacteria in foods. The combination of L. reuteri DPC16 and CA was assessed for its inhibitory effect on L. monocytogenes using DPC16-SCS and the fermentative supernatant of L. reuteri DPC16 from a glycerol-water solution (DPC16-GFS). The results showed that both of these supernatants, at 25 AU/mL, in combination with CA (60% CO2:40% N2) had a combined inhibitory effect on L. monocytogenes which could not be achieved by any one of the individual factors alone. Analysis of the levels of expression of some stress response genes of L. monocytogenes, after growth in the presence of L. reuteri DPC16 supernatant and/or CA, showed that the expression of some genes was affected including genes betL, gbuA and opuCA responsible for osmosis adaptation and genes gadA, gadB and gadC responsible for acid tolerance. Induction of gbuA, gadB and gadC by the culture supernatant suggests activation of osmotic and acid adaptation and that these genes play a major role in the culture supernatant-induced stresses. An investigation was also carried out to determine if the changes in gene expression conferred a cross-protection to heat. The result showed that the survival of L. monocytogenes grown in the presence of the culture supernatant and CA was significantly increased after exposure to heat treatment at 56oC, suggesting that a cross-protection to thermal stress had been induced. Based on these findings it is proposed that a comprehensive novel strategy incorporating both L. reuteri DPC16 or its fermentative products and a modified atmosphere rich in CO2 could be developed to potentially control foodborne pathogens in food products.