Thermal resistance of Bacillus stearothermophilus spores in different heating systems containing some approved food additives

M. LÓPEZ, M. MAZAS, I. GONZÁLEZ, J. GONZALEZ AND A. BERNARDO. 1996. The effects of different heating systems on the heat resistance of Bacillus stearothermophilus spores (ATCC 7953, 12980, 15951 and 15952) were investigated. Spores were heated in distilled water, Sorensen buffer (0.18 moll⁻¹), McIlvaine buffer (0.0025-0.18 moll⁻¹), and several solutions containing sodium chloride (0.0612%), sodium nitrite (125 ppm), potassium sorbate (0.1%) and sodium benzoate (0.1%) over a wide range of temperatures (115-140°C). D-values obtained for McIlvaine and Sorensen buffers, at the same molarities, were not significantly different (P > 0.05), but decimal reduction times increased as phosphate concentrations in the solutions decreased. The concentrations, in which statistically significant differences (P < 0.05) were obtained, varied among strains. Among the additives assayed, only sodium chloride reduced heat resistance, being effective at concentrations as low as 0.06%. The z-values calculated in this study ranged from 6.99 to 8.40 with a mean value of 7.60±0.45. Although z-values observed for salt and buffers (180 moll⁻¹) were slightly higher than those obtained in the other conditions assayed, the difference was not statistically significant (P > 0.05).     

The lack of identity between Bacillus stearothermophilus and Bacillus thermoglucosidasius in serological patterns of exo-oligo-1,6-glucosidase and exo-α-1,4-glucosidase

Summary Among 16 Bacillus stearothermophilus strains, 11 strains (ATCC 7953, ATCC 10149, ATCC 12976, ATCC 12978, ATCC 12980, ATCC 15951, ATCC 21365, IAM 11001, IAM 11004, IAM 11062 and IFO 12550) produced a protein reactable on double immuno-diffusion with the antiserum against Bacillus thermoglucosidasius KP 1006 (DSM 2542) exo-oligo-1,6-glucosidase (dextrin 6-glucanohydrolase, EC.3.2.1.10). However, these antigens in part shared their antigenic determinants. In addition to an exo-oligo-1,6-glucosidase, 6 B. thermoglucosidasius strains [KP 1006, KP 1012, KP 1013, KP 1014, KP 1019 and KP 1022 (DSM 2543)] formed a protein cross-reacted with the antiserum against B. stearothermophilus ATCC 12016 exo--1,4-glucosidase (-d-glucoside glucohydrolase, EC.3.2.1.20). These two antigens showed, however, a partial coincidence in their antigenic determinant groups. Of 16 B. stearothermophilus strains, 3 strains (ATCC 8005, ATCC 12016 and ATCC 15952) produced a protein immunologically compatible with the -1,4-glucosidase, while 4 strains (ATCC 12979, ATCC 12980, ATCC 15951 and IAM 11001) made the other protein which showed certain differences partly from this enzyme in its antigenic groups. No protein precipitated with the anti--1,4-glucosidase occurred in the remaining 9 B. stearothermophilus strains (ATCC 7953, ATCC 10149, ATCC 12976, ATCC 12977, ATCC 12978, ATCC 21365, IAM 11004, IAM 11062 and IFO 12550). These data indicate no serological identity between two thermophilic Bacillus species in their glucosidase patterns.     

Comparison of anti-Vibrio activities of potassium sorbate, sodium benzoate, and glycerol and sucrose esters of fatty acids.

The effects of fatty acids and their glycerol and sucrose esters, potassium sorbate, and sodium benzoate on growth of Vibrio parahaemolyticus in laboratory media at pH 6.7 were evaluated. The minimum concentrations at which inhibition by esters of glycerol could be detected were lowest for monolaurin (5 microgram/ml) and monocaprin (40 microgram/ml); these concentrations were lower than those observed for inhibition by lauric and capric acids, respectively. Inhibitory action of sucrose caprylate was detected at 40 microgram/ml, whereas sucrose caprate was effective at 100 microgram/ml; sucrose esters of lauric, myristic, and palmitic acids were ineffective at 100 microgram/ml. Potassium sorbate and sodium benzoate inhibited growth at concentrations as low as 30 and 300 microgram/ml, respectively, and enhanced the rate of thermal inactivation of V. parahaemolyticus at slightly higher concentrations. Fatty acid esters of glycerol and sucrose offer potential as perservatives for slightly acid or alkaline low-fat foods which do not lend themselves to the full antimicrobial action of traditional food preservatives such as potassium sorbate and sodium benzoate.     

Attributes of fresh gilt-head seabream (Sparus aurata) fillets treated with potassium sorbate, sodium gluconate and stored under a modified atmosphere at 0±1°C

The development of microbial populations on fillets of Mediterranean gilt-head sea bream ( Sparus aurata ) treated with potassium sorbate, sodium gluconate or a combination of both and stored under a modified atmosphere (MA) of 40% CO₂, 30% O₂ and 30% N₂ at 0±1 °C for about 30 d was studied. The pH of aqueous solutions of the preservatives was adjusted to 6·0 with HCl. The preservatives were applied by dipping. The use of sorbate plus gluconate was more effective than sorbate alone. Gluconate had a positive effect on the growth of Gram-positive micro-organisms. Changes in the concentrations of glucose, glucose-6-phosphate, ammonia, acetic acid, trimethylamine-nitrogen and sorbate were also monitored.     

Evaluation of Inhibitory Effect of Organic and Inorganic Salts Against Ilyonectria liriodendri,The Causal Agent of Root Rot Disease of Kiwifruit

This study evaluated efficacy of 42 organic and inorganic salts as possible alternatives to synthetic fungicides for the control of Ilyonectria root rot of kiwifruit. Preliminary in vitro tests showed that ammonium bicarbonate, ammonium carbonate, potassium benzoate, potassium sorbate, sodium benzoate and sodium metabisulphite at 2% completely inhibited mycelial growth of the fungus. No significant differences were observed among these salts and disodium EDTA (P ≤ 0.05). However, the ED₅₀, minimum inhibition concentration (MIC), and minimum fungicidal concentration (MFC) values indicated that sodium metabisulphite was more toxic to Ilyonectria liriodendri than these other six salts. Soil bioassays showed that sodium metabisulphite, sodium benzoate and potassium sorbate at 0.25% completely inhibited mycelial growth of the fungus, whereas potassium benzoate reduced the mycelial growth of fungus by 90.30%; however, the differences in inhibitory effects were statistically insignificant (P ≤ 0.05). Moreover, there was no significant difference between 0.1% sodium metabisulphite and 0.5% ammonium carbonate, 0.75% ammonium bicarbonate and 1.5–2.0% disodium EDTA (P ≤ 0.05). Unlike disodium EDTA, complete inhibitory was observed with ammonium carbonate and ammonium bicarbonate at higher concentrations. However, in root bioassays, applications of 2% ammonium bicarbonate, 1.5% ammonium carbonate and 2% disodium EDTA were phytotoxic to kiwifruit seedlings, but 0.25% four other salts were neither phytotoxic to kiwifruit seedlings nor did it adversely affect root length, root fresh weight and root dry weight of seedling. This study also showed I. liriodendri to be capable of growth in both acidic and basic environments. However, while the fungus showed uninhibited growth at pH values of 5–11, growth decreased significantly at both higher and lower pH values (P ≤ 0.05) and was completely inhibited at pH 12.     

Multiple modes of inhibition of spore germination and outgrowth by reduced pH and sorbate

Germination and outgrowth of three strains of Clostridium botulinum in PYEG medium were measured by phase contrast microscopy. Reduction in pH from 7 to 5·5 completely inhibited germination of strain 12885A, reduced the extent of germination of strain 62A and had no effect on the extent of germination of strain 53B. At pH 5·5, 225 mg/1 of undissociated sorbic acid had no effect on the germination of strain 53B, while at pH 6·5, 225 mg/1 of undissociated sorbic acid completely inhibited germination of strains 62A and 12885A. Outgrowth of germinated spores of strains 62A and 53B was not inhibited at pH 5·5, but the addition of sorbate (225 mg/1 undissociated sorbic acid) completely inhibited outgrowth. Sorbate inhibited germination of Cl. botulinum and Bacillus cereus spores triggered to germinate by amino acids. Inhibition occurred after germinant binding, as measured by commitment to germinate.     

Effect of Food Additives and Irradiation on Survival of Salmonella in Oysters

Salmonella typhimurium and S. enteritidis were inoculated into blended oysters, both raw and autoclaved. The oysters were also treated with sodium benzoate (0.1%) or potassium sorbate (0.1%), and irradiated (0.1 Mrad). In both non-irradiated and irradiated samples, greater numbers of Salmonella were recovered after storage at 7 C in the presence of sodium benzoate or potassium sorbate. The results of the autoclaved samples and studies in buffer indicated that this effect was not due to the reduction of competition from the natural flora when the additives were present.     

Effects of sporulation pH on the heat resistance and the sporulation of Bacillus cereus

Spores of Bacillus cereus ATCC 7004, 4342 and 9818 were obtained in nutrient agar at several pH from 5·9 to 8·3. The optimum pH for sporulation was around 7, but good production of spores was obtained in the range 6·5–8·3. With all three strains, D -values clearly dropped with sporulation pH, decreasing by about 65% per pH unit. z -Values were not significantly modified ( P > 0·05) by this factor. Mean z -values of 7·13 °C ± 0·16 for strain 7004, 7·67 °C ± 0·04 for 4342 and 8·80 °C ± 0·64 for 9818 were obtained.     

Combined effects of solutes and food preservatives on rates of inactivation of and colony formation by heated spores and vegetative cells of molds.

The combined and independent effects of sucrose, sodium chloride, potassium sorbate, and sodium benzoate on heat inactivation of conidia of Aspergillus flavus and Penicillium puberulum, ascospores of Byssochlamys nivea, and vegetative cells of Geotrichum candidum were studied. In addition, the effects of solutes and preservatives on colony formation by unheated and heated conidia of A. flavus were evaluated. Increased concentrations of sucrose were accompanied by increased tolerance to heat by A. flavus, B. nivea, and G. candidum. Low concentrations (3 and 6%) of sodium chloride protected A. flavus and G. candidum, whereas up to 12% sodium chloride protected B. nivea, but had little effect on the heat stability of P. puberulum. Potassium sorbate and sodium benzoate acted synergistically with heat to inactivate all four molds. At the same concentration, the two preservatives had varied degrees of effectiveness on molds and were influenced by the type of solute in the heating menstrua. Heated conidia of A. flavus had increased sensitivity to preservatives and reduced water activity, whether achieved by the presence of sucrose or sodium chloride, thus demonstrating heat-induced injury. At the same concentration, potassium sorbate was clearly more inhibitory than was sodium benzoate to colony formation by A. flavus, and the presence of sucrose and sodium chloride enhanced this inhibition.     

蕨类植物抗菌作用的初步研究

本文利用圆纸片法对十二种蕨类植物抗菌作用进行了研究,发现海金沙、凤尾蕨、抱石莲、兖州卷柏、乌韭、紫萁、翠云草等有较强的抗菌作用,在供试植物中,以海金沙抗菌活性最强.海金沙水提液和醇提液对金黄色葡萄球菌的最小抑制浓度(MIC)分别为10.、7.5%将海金沙、凤尾蕨、抱石莲三种植物等量组成复方,对该复方的水提液及醇提液与常用的化工合成防腐剂苯甲酸钠、山梨酸钾的防腐性能进行了比较.     

Destruction of Bacillus licheniformis spores by microwave irradiation

Aims:  To investigate the sporicidal mechanisms of microwave irradiation on Bacillus licheniformis spores.
Methods and Results:  We measured spore viability and the release of DNA and proteins, and performed transmission electron microscopy (TEM). A microwave oven (0·5 kW) was modified to output power at 2·0 kW, which allowed a shorter sterilization cycle. A 2·0 kW microwave treatment at the boiling temperature for 1 min did not kill all spores, but killed most spores. The spore inactivation rate was faster than that of boiling and 0·5 kW microwave oven. In contrast to boiling and 0·5 kW microwave treatments, the 2·0 kW microwave resulted in significant leakage of proteins and DNA from spores due to injury to the spore structure. TEM revealed that 2·0 kW microwave irradiation affected spore cortex hydrolysis and swelling, and ruptured the spore coat and inner membrane.
Conclusions:  These results suggest that 2·0 kW microwave irradiation ruptures the spore coat and inner membrane, and is significantly different from boiling.
Significance and Impact of the Study:  This study provides information on the sporicidal mechanisms of microwave irradiation on B. licheniformis spores.     

Synergy between <Emphasis Type="Italic">Salvia officinalis</Emphasis> L. and some preservatives

The aim of the present study is to investigate the antibacterial activity of Salvia officinalis L. aqueous extracts and its synergistic action with preservatives sodium nitrite, sodium benzoate and potassium sorbate in vitro against selected food spoiling bacteria. Synergy was assessed by the checkerboard assay method and quantitatively represented by the FIC index. Synergistic action was established for aqueous extract/ sodium benzoate, aqueous extract/ potassium sorbate, aqueous extract/ sodium nitrite combinations. Synergy was detected in relation to: Agrobacterium tumefaciens, Bacillus subtilis and Proteus sp. Synergy was established at plant extract and preservative concentrations corresponding up to 1/8 MIC values.     

The effect of recovery conditions on the apparent heat resistance of Bacillus cereus spores

The effect of recovery media and incubation temperature on the apparent heat resistance of three ATCC strains (4342, 7004 and 9818) of Bacillus cereus spores were studied. Nutrient Agar (NA), Tryptic Soy Agar (TSA), Plate Count Agar (PCA) and Milk Agar (MA) as the media and temperatures in the range of 15–40°C were used to recover heated spores. Higher counts of heat injured spores were obtained on PCA and NA. The optimum subculture temperature was about 5°C below the optimum temperature for unheated spores. No significant differences in heat resistance were observed with the different recovery conditions except for strains 4342 and 9818 when MA was used as plating medium.
Large differences in D -values were found among the strains ( D ₁₀₀=0·28 min for 7004; D ₁₀₀=0·99 min for 4342; D ₁₀₀= 4·57 min for 9818). The 7004 strain showed a sub-population with a greater heat resistance. The z values obtained for the three strains studied under the different recovery conditions were similar (7·64°C 0·25).     

Effect of potassium sorbate,sodium benzoate,and sodium nitrite on biosynthesis of cyclopiazonic and mycophenolic acids and citrinin by fungi of the <Emphasis Type="Italic">Penicillium</Emphasis> genus

The effect of potassium sorbate, sodium benzoate, and sodium nitrite used as preservatives in the food industry in the production of such mytotoxins as citrinin cyclopiazonic and mycophenolic acids by the contaminating fungi Penicillium citrinum, P. commune, and P. brevicompactum, respectively, was investigated. It was shown that the effect of preservatives used at concentrations relevant to the food industry on the synthesis of mycotoxins depended on the species-specific biochemical and physiological features of the cultures. The growth of P. brevicompactum was inhibited to the highest degree by sodium nitrite and potassium sorbate, and the growth of P. commune was so inhibited by sodium benzoate. It was established that the introduction of 0.015% sodium nitrite into the medium resulted in 1.3- and 1.4-fold reductions of the production of citrinin and mycophenolic acid, respectively, while the production of cyclopiazonic acid did not change in comparison with the control. The introduction of 0.015% sodium benzoate caused a more than 1.5-fold increase of the concentration of citrinin, cyclopiazonic, and mycophenolic acids, and the addition of 0.02% potassium sorbate increased the production of cyclopiazonic and mycophenolic acids by 1.7 and 2.6 times, respectively.     

Inactivation effect of X-ray treatments on Cronobacter species (Enterobacter sakazakii) in tryptic soy broth, skim milk, low-fat milk and whole-fat milk*

Aims:  To determine the inactivation effect of X-ray treatments on Cronobacter ( E. sakazakii ) in tryptic soy broth (TSB), skim milk (0% fat), low-fat milk (1% and 2%) and whole-fat milk (3·5%).
Methods and Results:  X-rays were produced using the RS 2400 generator system (Rad Source Technologies Inc.). Cronobacter (in TSB), inoculated skim milk (0% fat), low-fat milk (1% and 2% fat) and whole-fat milk (3·5% fat) were treated with 0·0, 0·1, 0·5, 0·75, 1·0, 2·0, 3·0, 4·0, 5·0 and 6·0 kGy X-ray doses. Surviving bacteria in the TSB and inoculated milk, before and after treatment, were enumerated using plating method onto trypticase soy agar. Greater than 7·0-log CFU reduction in Cronobacter population was observed with 4·0, 5·0, 6·0, 6·0 and 6·0 kGy X-ray in the TSB, skim milk, 1% fat milk, 2% fat milk and 3·5% fat milk, respectively.
Conclusions:  Treatment with X-rays significantly ( P  <   0·05) reduced Cronobacter to less than detectable limits (<1 log CFU ml⁻¹) in skim milk at 5·0 kGy and milk with 1% fat content and greater at 6·0 kGy dose levels. The D-value for Cronobacter in TSB was significantly ( P  <   0·05) lower than those in milk samples.
Significance and Impact of the Study:  Treatment with X-rays could be an effective and safe alternative technology to control pathogenic bacteria ( Cronobacter ) in the dairy industry.     

Polyaluminum chloride-enhanced concentration efficiency of poliovirus and f2 phage from sewage water

Aims:  To enhance the recovery of f₂ bacteriophage and poliovirus by an established method based on the adsorption to and elution from positively-charged Al(OH)₃-treated silica gel.
Methods and Results:  Polyaluminum Chloride (PAC) was added to water samples to neutralize the negatively charged materials, which can reduce virus recovery by providing a competing adsorption mode on the media surface. Using this improved process (PAC 30 mg l⁻¹, pH 6·5, temperature 20∼30°C), the recoveries of Poliovirus I and f₂ from small-volume sewage (100 ml) were 110·76 ± 36·0% and 92·06 ± 8·65%, respectively ( P  < 0·05 vs. traditional methods). Recovery from a 20-L volume of sewage averaged 85·65 ± 4·43% for f₂ and 88·73 ± 9·76% for poliovirus, significantly higher than the recoveries in the traditional methods ( P  < 0·05).
Conclusions:  PAC could enhance concentration efficiency of poliovirus and f₂ phage from sewage water.
Significance and Impact of the Study:  This method should significantly improve the recovery of viruses from sewage.     

Use of organic acids and salts to control postharvest diseases of lemon fruits in Egypt

Abstract

Postharvest diseases caused by Geotricum candidum (sour rot), Penicillium digitatum (green mould), and P. italicum (blue mould) are the most important negative factors affecting handling and marketing of citrus fruits in Egypt. The effect of organic acids (ascorbic, benzoic, citric and sorbic) as well as organic salts (potassium sorbate and sodium benzoate) were evaluated on the growth of causal agents and their disease incidence under in vitro and in vivo conditions. Complete inhibition was observed in the linear growth of all tested fungi when exposed to benzoic, citric and sorbic organic acids at concentrations of 4% and 2% of either sodium benzoate or potassium sorbate, respectively. Minimizing the tested concentration of organic acid down to 2%, the tested fungi fluctuated in their response such that only benzoic and sorbic acids could completely inhibit the growth of either P. digitatum or P. italicum only. Different organic acids and salts showed various levels of either protective or therapeutic effect for coated lemon fruits against mould infection whatever the time of their artificial inoculation under in vivo conditions. All treated fruits showed reduction in sour rot and green and blue mould diseases when compared with untreated fruits. Complete inhibition of mould incidence was obtained in coated lemon fruits with 4% of water or wax mixtures of sodium benzoate and potassium sorbate 24 hours before inoculation. Also, high reduction in mould incidence was observed in lemon fruits coated with the same concentration at 48 hours after inoculation under the same conditions. On the other hand, the tested organic acids showed a lesser effect on mould incidence. Moreover, they were more effective against mould incidence when dissolved in water than wax, that only 4% of water mixture of sorbic and benzoic acids showed 100% protection against mould incidence. Furthermore, the severity of infection records followed the same trend. The present findings demonstrate that potassium sorbate and sodium benzoate have potential as environmentally friendly products, nontoxic postharvest fungicides against sour rot, green and blue mould incidence of stored citrus fruits and could be suggested for commercial use in packing-houses in consideration to their wide consumption as safely food preservatives.     

Modeling yeast spoilage in cold-filled ready-to-drink beverages with Saccharomyces cerevisiae, Zygosaccharomyces bailii, and Candida lipolytica

Mathematical models were developed to predict the probability of yeast spoilage of cold-filled ready-to-drink beverages as a function of beverage formulation. A Box-Behnken experimental design included five variables, each at three levels: pH (2.8, 3.3, and 3.8), titratable acidity (0.20, 0.40, and 0.60%), sugar content (8.0, 12.0, and 16.0 degrees Brix), sodium benzoate concentration (100, 225, and 350 ppm), and potassium sorbate concentration (100, 225, and 350 ppm). Duplicate samples were inoculated with a yeast cocktail (100 microl/50 ml) consisting of equal proportions of Saccharomyces cerevisiae, Zygosaccharomyces bailii, and Candida lipolytica (approximately 5.0 x 10(4) CFU/ml each). The inoculated samples were plated on malt extract agar after 0, 1, 2, 4, 6, and 8 weeks. Logistic regression was used to create the predictive models. The pH and sodium benzoate and potassium sorbate concentrations were found to be significant factors controlling the probability of yeast growth. Interaction terms for pH and each preservative were also significant in the predictive model. Neither the titratable acidity nor the sugar content of the model beverages was a significant predictor of yeast growth in the ranges tested.     

Effectiveness of various food preservatives in controlling the outgrowth of Byssochlamys nivea ascospores

Potassium sorbate, sodium benzoate, sulfur dioxide, and diethylpyrocarbonate (DEPC) were tested for their effectiveness in preventing the outgrowth ofByssochlamys nivea Westling ascospores. Sulfur dioxide was the most inhibitory of the test antimycotics, complete inhibition of colony formation occurring in acidified (pH 3.5) potato dextrose agar containing 50 ppm of the preservative. Complete inhibition ofB. nivea ascospore outgrowth in grape juice stored for 60 days was noted in the presence of 300 ppm sulfur dioxide, 400 ppm potassium sorbate, and 600 ppm DEPC. Growth was observed in grape juice containing 1000 ppm sodium benzoate. The presence of up to 100 ppm potassium sorbate in grape juice during heat activation appears to have a stimulatory effect on breaking dormancy, while the other test preservatives at this concentration decrease the heat resistance ofB. nivea ascospores. The time elapsed between heat shock and exposure to DEPC or sodium benzoate is critical with respect to the sensitivity of ascospores to these preservatives.     

Effects of potassium sorbate and other antibotulinal agents on germination and outgrowth of Clostridium botulinum type E spores in microcultures.

The effects of potassium sorbate, sodium hypophosphite, sodium tripolyphosphate, sodium nitrite, and linoleic acid on the germination and outgrowth of Clostridium botulinum type E spores were studied in microcultures. At pH 5.8 to 6.0 in liver veal agar, the germination rate was decreased to nearly zero with 1.0, 1.5, or 2.0% sorbate. At pH 7.0 t 7.2, these levels of sorbate afforded germination and outgrowth of abnormally shaped cells that were defective in cell division. At the high pH range, 0.5 or 1.0% hypophosphite had effects similar to those of sorbate. The use of 0.05% sodium nitrite with sorbate enhanced the lysis of outgrowing cells at pH 7.2 or lower. Emergence and elongation were inhibited by 0.05% linoleic acid with or without 1.0% sorbate at pH 7.0 to 7.2. The addition of 0.5% tripolyphosphate to media containing 1.5% sorbate at pH 7.1 prevented normal cell growth to an extent greater than with sorbate alone.