A biological indicator (Bl) for use in thermal or chemical sterilization processes consists of standardized bacterial spore preparations which are usually in the form either of suspensions or of spores dried on carriers. Bl is usually placed in dummy packs located at strategic sites in the sterilizer. After the sterilization process, the aqueous suspensions or spores on carriers are aseptically transferred to an appropriate culture medium, which is then incubated and periodically examined for signs of growth. Spores of Geobacillus stearothermophilus ATCC 7953 or 12980 are used for steam, formaldehyde, hydrogenperoxide, ozone sterilization monitoring, and these are incubated at 55°C. Spore of Bacillus atrophaeus ATCC 9372 is used for ethylene oxide and dry heat sterilization monitoring, and incubated at 30-35°C. When the paper strip type Bl is used, aseptic transfer is indispensable. Aseptic transfers can be avoided by the use of self-contained Bl where the spore strip and nutrient medium are present in the same device ready for mixing after use. This Bl is, however, not recommended for use in a validation study. Resistance of Bl is adjudged from the spore destruction curve obtained upon exposure to the sterilization process, as well as the recommended Bl spores and their decimal reduction times. Great care must be taken in the preparation and storage to ensure a reproducible response to sterilization processes to assure that Bl is a reliable monitor for steriliza tion validation. One of the drawbacks of Bl is that a long period of incubation is required to confirm a satisfactory sterilization process, which imposes an undesirable delay on the release of the product. To shorten the incubation period is now being discussed at ISO TC 198, but as sterilized Bls are injured spores, a short cultivation time may result in an inaccurate result.

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An aromatic polysulfone (PS) consists of a 4, 4'-diol aromatic compound and 4, 4'-dichiorodiphenyl sulfone. As 4, 4'-diol aromatic compounds, bisphenol A, p-dihydroxy benzene, 4, 4'-diphenol methane and p, p'-diphenol (bisphenol) were used for the fabrication of new PSs. These PSs were tested to see which PS would show the greatest tolerance to gamma ray irradiation. Sulfur dioxide (SO₂) production from irradiated PSs was used as an indicator to evaluate their stability. The bisphenol-PS was the most tolerant to the gamma ray irradiation and had the smallest production of SO₂. This indicated that a PS free from bisphenol A was obtainable. A fabricated PS tolerates to gamma ray irradiation and is safer to people due to the absence of bisphenol A, an endocrine disrupter. The rate of decrease of tensile strength correlated well with the order of radiation resistance. The fracture toughness of bisphenol A-PS decreased with irradiation doses, but bisphenol-PS almost maintained its original ductility.

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A resistant strain of Escherichia coil IFO12713, resistant against the quaternary ammonium compound (QAC), N-dodecylpyridinium iodide (P-12), was developed by using a standard broth dilution method and physiologically characterized by comparison to a sensitive strain. The cell surface hydrophobicity cultivated in the resistance-acquiring process gradually decreased and the lipopolysaccharides in the outer membrane increased. This result indicated that the cell surface of the resistant strain was more hydrophobic than that of the sensitive strain. One of the resistant strains isolated from the process also had resistance to other antibiotics and organic solvents. Rapid ethidium efflux was observed in the resistant strain compared to the sensitive strain. To elucidate the mechanism of resistance on a molecular level, protein analysis was performed with 2D-PAGE for the entire cellular fraction, and SDS-PAGE for the outer or inner membrane fractions. It was seen that 18 spots and bands of proteins specifically increased in the resistant strain. From the physiological characterization and the protein analysis, it was suggested that the QAC-resistant strain acquired its resistance by responding to antimicrobial stresses, stabilizing the outer membrane to control the adsorption/binding reaction between the cell and agent, and activating the efflux pump against the antimicrobial agents.

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An automated fluorescence microscopic method developed for the measurement of viable bacterial counts was applied to the detection and enumeration of viable bacteria in milk. In the automated analysis of viable bacteria, bacterial cells were recovered from milk after the treatments with EDTA and Triton X-100, stained with 6-CFDA, and the stained cells were counted with the automated microscopic method. Salmonella Enteritidis expressing the enhanced green fluorescent protein (S. Enteritidis-EGFP) was used to elucidate the relationship between the fluorescent bacterial counts by the automated microscopic method and the viable counts by the conventional plating method. A high correlation coefficient (r²=0.97) was obtained for the relationship between the counts by the microscopic method and those by the plating method. When S. Enteritidis-EGFP was recovered from sterile milk after treatments with EDTA and Triton X-100, the correlation coefficient was calculated to be r²=0.91. Viable counts by the plating method and the counts of 6-CFDA stained bacterial cells by the automated microscopic method were determined for various raw milk samples. Thirty-seven raw milk samples were measured and the correlation coefficient was calculated to be r²=0.73.

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We assayed in vitro the biological activities of caffeic acid phenethyl esters that had been enzymatically synthesized from 5-chlorogenic acid and phenethyl alcohol. Caffeic acid phenethyl esters showed antioxidant and hyaluronidase-inhibitory activities similar to those of chlorogenic acid and caffeic acid, and markedly higher antibacterial, antimutagenic, and antiviral activities than chlorogenic acid. The antimicrobial activities of caffeic acid phenylethyl esters against Staphylococcus aureus, Bacillus subtiiis, and Pseudomonas aeruginosa (MIC; 0.22-0.44, 0.44-0.88, and 0.44-0.88 mM, respectively) were higher than those of 5-chlorogenic acid and caffeic acid. 2-caffeic acid phenylethyl ester showed potent antimutagenic activity against quinoline compounds, aromatic amines, and heterocyclic compounds. In addition, 2-caffeic acid phenylethyl ester at concentrations of less than 35 μM promoted the proliferation of normal human pulmonary fibroblasts (WI-38 cells) in culture, but inhibited the proliferation of virus-transformed fibroblasts (WI-38 VA cells; IC₅₀ of 60 μM). 1-and 2-Caffeic acid phenylethyl esters at a concentration of 8.8 μM inhibited the growth of type A influenza virus by 95% and 96%, respectively, and that of type B influenza virus by 92% and 94%, respectively. These results indicate that caffeic acid phenylethyl esters have higher antimutagenic and anti-influenza virus activities than chlorogenic acid and caffeic acid.

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Persimmon tannin is especially known well as a fruit polyphenol. It inhibited the growth of streptococci of the mutans group. It also exhibited a strong inhibitory effect on glucosyltransferase-1(GTase-1) activity. The sucrose-dependent adherence of the bacterial cells was examined with persimmon tannin in vitro. The tannin inhibited the production of adherent materials at the lowest concentration among the polyphenols tested. Persimmon tannin exhibited an inhibitory effect on cariogenic factors at the steps of tooth-decaying bacterial growth, insoluble glucan synthesis, and dental plaque formation, and is expected to be a used for the prevention of dental caries.

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