Sourdough

Rye bread, several wheat bread types, and some sweet baked goods have traditionally been fermented with sourdough. Besides improving the flavor and texture of the bread, this mixture of lactic acid bacteria and yeast increases the shelf lives of these products (38).

The antimicrobial effect oflactic acid bacteria has been assigned to both the lactic and acetic acids produced during fermentation (39,40); newer studies, however, have shown that lactic acid does not have an antimicrobial effect besides the pH-lowering effect (41-43). The antifungal effect of the dough seems to be more directly related to the acetic acid content (44). The antifungal spectrum oflactic acid bacteria varies, and antifungal effect has mainly been found in obligate heterofermentative Lactobacillus spp. Of these, Lactobacillus sanfranciscensis has shown the largest spectrum of antifungal activity (41). It produced a mixture of acetic, caproic, formic, butyric, and n-valeric acids, which synergistically inhibited species of Fusarium, Penicillium, Aspergillus, and Monilia. Caproic acid played a key role in inhibiting mold growth (41).

Lactobacillus sanfranciscensis is often isolated from sourdoughs in association with L. plantarum. This species has also been shown to produce antimicrobial compounds (43). These compounds corresponded to benzoic acid, 5-methyl-2,4-imidazolidinedione, tetrahy-dro-4-hydroxy-4-methyl-2H-pyran-2-one, and 3-(2-methylpropyl)-2,5-piperazinedione and were inhibitory to Pantoea agglomerans and also inhibited the fungus Fusarium avenaceum to some extent.

Another L. plantarum strain (21B) isolated from sourdough in Italy showed a very broad spectrum of activity against the most important bread-associated fungi (16). Spoilage by A. niger occurred after 2 days in breads started with Saccharomyces cerevisiae alone or with S. cerevisiae and the acidifying lactic acid bacterium Lactobacillus brevis, whereas fungal growth was not evident before day 7 on bread started with S. cerevisiae and L. plantarum 21B. L. plantarum 21B was found to produce a range of metabolites, of which phenyllactic acid was found to play a key role in inhibiting fungal growth.

Some sourdough cultures have also been proven effective against rope-forming B. subtilis and B. licheniformis spores. Rosenquist and Hansen (45) found that B. subtilis growth could be prevented by 0,1% v/w propionic or acetic acid or 15% sourdough fermented with L. plantarum, Lact. brevis, and Lact. meltaromicus. Katina and coworkers (46) found an almost similar effect of sourdough fermented with L. palantarum and Peciococcuspentosaceus. Here 20-30% sourdough could hinder rope formation.

C. Natural Preservatives

The antimicrobial properties of herbs and spices have been recognized and used for food preservation and medical treatments since ancient times (47,48). Scientific reporting on natural antimicrobial agents also dates back more than a century; Chamberlain reported in 1887 the action of essential oil vapors on anthrax spores, as cited by Maruzzella and Sicurella (49). A renewed interest in 'natural preservation' seems to be stimulated by present food safety concerns, growing problems with microbial resistance, and a rise in production of minimal processed food joined with 'green'-image policies of food industries.

Numerous studies have documented the antifungal (50-55) and antibacterial (56-58) effect of plant essential oils. Examination of indigenous and local herbs and plant material have been reported from all over the world, e.g., India (59) Australia (60), Argentina (61) and Finland (62).

Screening experiments with 13-52 essential oils and major active components against 5-25 microorganisms (63-67) have reported thyme, clove, cinnamon, bay, oregano, garlic, and lemongrass to be some of the best broad-spectra candidates for inhibition of food-borne pathogens and spoilage organisms. By comparing studies that have used different methodologies, it is clear that the minimal inhibitory concentrations (MIC) are highly dependent on the test method. Suhr and Nielsen (68) examined the application method and found large differences between the antimicrobial effects, whether they were added to the media or added as a volatile to the packaging atmosphere. Figure 1 is a loading plot from a partial least squares (PLS) regression of degree of inhibition of fungal growth as affected by concentration and application method of a range of essential oils. Oils consisting of smaller volatile compounds were most efficient when applied through the gas phase. Mustard (1 ppm) and lemongrass completely inhibited growth of all fungi in the volatile system (marked with suffix V) for 14 days. Orange, thyme, sage, and rosemary A also strongly inhibited growth, but not equally well, all fungi.

Larger phenolic compounds worked better in direct contact (situated close to the upper right corner in Fig. 1). Thyme, clove and cinnamon almost completely inhibited growth for up to 14 days. Mustard (250 ppm) and to some degree lemongrass were also

Figure 1 The inhibitory effect of some essential oils on the most important bread spoilage fungi: Aspergillusflavus = Afla; Erotium repens = Erep; Endomycesfibuliger = Efib; Penicillium roqueforti = Proq; P. corylophilum = Pcor as affected by application method. All oils were added in same amount in gas phase or in media (approx. 125 and 250 ppm) except mustard oil, of which only 1 and 2 ppm (aI/L) were added to the volatile gas phase (V). Oils situated closest to the upper left corner and the fungi with suffix V work well in a volatile system, whereas oils situated closest to the upper right corner work best when added to the media.

Figure 1 The inhibitory effect of some essential oils on the most important bread spoilage fungi: Aspergillusflavus = Afla; Erotium repens = Erep; Endomycesfibuliger = Efib; Penicillium roqueforti = Proq; P. corylophilum = Pcor as affected by application method. All oils were added in same amount in gas phase or in media (approx. 125 and 250 ppm) except mustard oil, of which only 1 and 2 ppm (aI/L) were added to the volatile gas phase (V). Oils situated closest to the upper left corner and the fungi with suffix V work well in a volatile system, whereas oils situated closest to the upper right corner work best when added to the media.

highly effective but the effect levelled off with time. These results indicate that mustard oil is inactivated when in contact with the media, because more than 250 times more had to be added to get the same effect in the media as in air.

For inhibition of mold growth on bread, smaller volatile compounds, such as allyl isothiocyanate and limonene, were most efficient when applied through the gas phase. Small volatile compounds are the most interesting for active packaging applications (see below) (69). Larger phenolic compounds, such as thymol and eugenol, were the best compounds for direct addition to the product.

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