• Journal of Microbiology and Biotechnology
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• v.19 no.12
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• pp.1590-1595
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• 2009

Alternative methods for controlling bacterial endospore contamination are desired in a range of industries and applications. Attention has recently turned to natural products, such as essential oils, which have sporicidal activity. In this study, a selection of essential oils was investigated to identify those with activity against Bacillus subtilis spores. Spores were exposed to 13 essential oils, and surviving spores were enumerated. Cardamom, tea tree, and juniper leaf oils were the most effective, reducing the number of viable spores by 3 logs at concentrations above 1%. Sporicidal activity was enhanced at high temperatures ($60^{\circ}C$) or longer exposure times (up to 1 week). Gas chromatography-mass spectrometry analysis identified the components of the active essential oils. However, none of the major oil components exhibited equivalent activity to the whole oils. The fact that oil components, either alone or in combination, did not show the same level of sporicidal activity as the complete oils suggested that minor components may be involved, or that these act synergistically with major components. Scanning electron microscopy was used to examine spores after exposure to essential oils and suggested that leakage of spore contents was the likely mode of sporicidal action. Our data have shown that essential oils exert sporicidal activity and may be useful in applications where bacterial spore reduction is desired.

• Journal of Food Hygiene and Safety
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• v.24 no.4
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• pp.312-317
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• 2009

Usually, bacterial spores are hundreds or thousands of times more resistant to chemical sanitizers than are vegetable bacteria. Consequently, it is hard to assess whether a commercial sterilant, containing hydrogen peroxide and peracetic acid as ingredients, has or does not have sporicidal activity under certain conditions using the National Standard Test Method for assessing bactericidal activity against Escherichia coli and Staphylococcus aureus. Hence we established alternative the standard test method and requirements to determine whether they are effective in showing at least reduction of $10^3$ in the number of Bacillus subtilis ATCC 6633 spores under the required test condition for evaluation of sporicidal activity including verification methodology. This standardized method has proved to be suitable for evaluating effectiveness of commercial sterilants and could be used as Standardization Test Method for assessing sporicidal activity.

• Food Science and Biotechnology
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• v.16 no.6
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• pp.1072-1077
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• 2007

Bacterial endospores, especially those of Bacillus and Clostridium genera, are the target of sterilization in various foods. We used Bacillus subtilis ATCC 6633 spores to screen novel antimicrobial substances against spores from medicinal plants. We collected 79 types of plant samples, comprising 42 types of herbs and spices and 37 types of medicinal plants used in traditional medicine in Korea and China. At a concentration of 1%(w/v), only 14 of the ethanol extracts exhibited antimicrobial activity against B. subtilis spores of at least 90%. Crude extracts of Torilis japonica, Gardenia jasminoides, Plantago asiatica, Fritllaria, and Arctium lappa showed particularly high sporicidal activities, reducing the spore count by about 99%. Consideration of several factors, including antimicrobial activity, extraction yields, and costs of raw materials, resulted in the selection of T. japonica, G. jasminoides, A. lappa, and Coriandrum sativum for the final screening of novel antimicrobial substances. Verification tests repeated 10 times over a 4-month period showed that the ethanol extract of T. japonica fruit reduced aerobic plate counts of B. subtilis spores the most, from $10^7$ to $10^4\;CFU/mL$ (99.9%) and with a standard deviation of 0.21%, indicating that this fruit is the most suitable for developing a novel antimicrobial substance for inactivating B. subtilis spores.

• Food Science and Biotechnology
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• v.18 no.5
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• pp.1301-1304
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• 2009

Macelignan is a bioactive compound isolated from nutmeg (Myristica fragrans Houtt.) which has been traditionally used for the food and pharmaceutical purposes. In this study, the activities of macelignan against vegetative cells and spores of Bacillus cereus were evaluated in vitro. Our results showed that the vegetative cells of B. cereus were significantly inhibited in growth by macelignan with minimum inhibitory concentration (MIC) of $4{\mu}g/mL$. The vegetative cells of B. cereus were completely killed with minimum bactericidal concentration (MBC) of $8{\mu}g/mL$ of macelignan. Killing time of macelignan against vegetative cells of B. cereus was very fast; endpoint of macelignan was reached after 4 hr of incubation at $4{\times}MIC$. Macelignan inactivated more than 3-log (99.9%) of spores/mL of B. cereus at the concentration of $100{\mu}g/mL$. Macelignan was found to be effective against vegetative cells and spores of B. cereus. These results suggest that macelignan might be good to be developed as a food preservative.

• Journal of Food Hygiene and Safety
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• v.29 no.3
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• pp.217-222
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• 2014

This study was performed to evaluate the sporicidal efficacy of a fumigation disinfectant containing 20% ortho-phenylphenol against Clostridium perfringens (C. perfringens) spores. In this research, efficacy test of fumigant against C. perfringens spores was carried out according to French standard NF T 72-281. C. perfringens spores working culture suspension number (N value), all the spore numbers on the carriers exposed with the fumigant (n1, n2, and n3), the number of bacterial spore suspensions by pour plate method (N1), the number of bacterial spore suspensions by filter membrane method (N2) and the mean number of bacterial spore recovered on the control-carriers (T value) were obtained from the preliminary test. In addition, the reduction number of C. perfringens spores exposed with the fumigant (d value) was calculated using T value, the mean number of bacterial spore in recovery solution (n'1) and the mean number of bacterial spore on carriers plated in agar (n'2). N value was $4.3{\times}10^7spores/mL$, and n1, n2, and n3 were higher than 0.5N1, 0.5N2 and 0.5N1, respectively. Additionally, T value was $4.9{\times}10^5spores/carrier$. In the sporicidal effect of the fumigant, the d value was 4.52log reduction. According to the French standard for the fumigant, the d value for the effective sporicidal fumigant should be over than 3log reduction. The results indicated that Fumagari $OPP^{(R)}$ had an efficient sporicidal activity against spores of C. perfringens, then the fumigant can be applied to disinfect food materials and kitchen appliances contaminated with bacterial spores.

• Food Science and Biotechnology
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• v.17 no.3
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• pp.603-607
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• 2008

Thirteen commercial antimicrobial products were examined to assess the sporicidal activity against Bacillus cereus spores at room temperature, 60 and $85^{\circ}C$. Neither the antimicrobials showed detectable antimicrobial activity against the B. cereus spores nor induced spore germination after the treatment at 0.5 or 1.0%(w/v, v/v) commercial antimicrobial agents at room temperature for 0.5 to 4 hr. However, when the antimicrobials such as chitosan, lactic acid, fermented pollen, grapefruit extract were applied with heat at $85^{\circ}C$ for 30 min, more than 1 log CFU/mL spores were additionally inactivated compared to only heat treatment without antimicrobials. Imposition of $60^{\circ}C$ to B. cereus spores with the higher concentration of 5.0%(v/v) lactic acid or 2.5%(w/v) thiamine dilaurylsulfate for the longer time incubation of 24 hr resulted in 3 log CFU/mL spore inactivation. This work showed that low concentrations of commercial antimicrobials by themselves did not inactivate B. cereus spores. However, when physical processes such as heat were combined together, antimicrobials showed a synergistic effect against B. cereus spores.