Recovery of nicotine-free proteins from tobacco leaves using phosphate buffer system under controlled conditions.

Establishment of an effective, high-throughput processing system to recover protein from tobacco with no nicotine contamination is essential and vital to the development of value-added, alternative applications for tobacco farmers. We have successfully developed a mechanism capable of processing up to 60 kg of tobacco leaves per hour with phosphate buffer (Na(2)HPO(4)-KH(2)PO(4)) simultaneously added to stabilize the protein as the plant was being disintegrated. The optimal processing parameters were identified, including the ratio of buffer to leaf (BLR) at 4.75 (w/w), buffer pH 7.85, and buffer concentration 0.085 mol/L, achieving a maximum yield of soluble protein at 12.85 mg/g fresh leaf. Acetone at -20 degrees C was the most effective among all methods investigated to remove nicotine from protein; however, it also drastically reduced the recovery rate of protein (63.3%). Ultrafiltration was only able to remove about 50% of the residual nicotine, although the protein recovery rate was high (94.7%). The residual nicotine content inherent in the recovered protein was completely removed by rinsing the protein with 85% phosphoric acid at pH 3.5 for three times with a protein recovery of 94.5%. The pilot-scale operation provides a solid foundation for further scale-up to industrial production of nicotine-free tobacco protein that could bring added value to tobacco for nonsmoking applications.

Invasiveness and intracellular growth of multidrug-resistant salmonella and other pathogens in Caco-2 cells.

The increase of multidrug-resistant pathogens of human and animal origins is a major public health concern. For a better understanding of the health consequences of multidrug-resistant bacteria transmitted from animal products to humans, the host interaction of zoonotic Salmonella isolates along with other pathogenic and commensal bacteria was evaluated using a human intestinal Caco-2 cell system. Multidrug-resistant S. Agona, S. Heidelberg, and S. Typhimurium possessed plasmid-mediated class 1 integrons. The S. Typhimurium DT104 isolate from ground beef showed the well-known genotypic and phenotypic resistance characteristics of the species, and contained the chromosomally located class 1 integron. Among the multidrug-resistant Salmonella isolates, the S. Heidelberg 219 had the highest invasion number at 1.0 x 10(4) CFU/mL, followed by the S. Typhimurium DT104 isolate at 7.7 x 10(3) CFU/mL. Listeria monocytogenes was the best performer among the tested species in invading the Caco-2 cell. Multidrug-resistant opportunistic pathogens Klebsiella pneumoniae and Pseudomonas aeruginosa were also able to invade the cells. The invasion of S. Heidelberg 219, S. Typhimurium DT104, L. monocytogenes, K. pneumoniae, and P. aeruginosa into the Caco-2 cells was not affected even in the presence of commensal E. coli. During the intracellular growth of S. Heidelberg 219, S. Typhimurium DT104, and L. monocytogenes, the bacterial counts increased 2 log cycles in 9 h in the Caco-2 cells. Therefore, these strains could rapidly proliferate after their invasion into the cells.

Expression of AmpC beta-lactamase in Enterobacter cloacae isolated from retail ground beef, cattle farm and processing facilities.

AIMS: To better understand antibiotic resistance of Enterobacter cloacae isolates originated from food animals, the phenotypic and genotypic resistance of Ent. cloacae isolates from retail ground beef, cattle farm, processing facilities and clinical settings were investigated. METHODS AND RESULTS: The ampC, ampD and ampR genes in the isolates were sequenced and analysed. beta-Lactamase activities and beta-lactamase profiles of the isolates were analysed by the enzymatic hydrolysis of nitrocefin and isoelectric focussing, respectively. The ampC gene of the Ent. cloacae isolate was cloned and transformed into Escherichia coli strains. The genomic DNA profiles of Ent. cloacae isolates were analysed by using pulse field gel electrophoresis (PFGE). Mutation at one residue (Val-54-->Ile) in the AmpR amino acid sequence was consistently found in Ent. cloacae isolates that were resistant to a broadspectrum of beta-lactam agents. The enzyme activity in the isolates was induced by cefoxitin. The pI (isoelectric point) of the enzymes produced by the test strains ranged from 8.4 to 8.9. Cloning of ampC gene of the Ent. cloacae isolate conferred the resistance to ampicillin, cephalothin and amoxicillin in recipient E. coli strains. One recipient of E. coli O157:H7 strain additionally acquired resistance to ceftiofur. The genomic analysis of Ent. cloacae isolates by PFGE showed that the isolates from various sources were genetically unrelated. CONCLUSIONS: The spread of diverse clones of AmpC-producing Ent. cloacae occurred in the ecosystem and retail products. SIGNIFICANCE AND IMPACT OF THE STUDY: Our findings suggested that AmpC-producing Ent. cloacae could be a contributor in spreading beta-lactamase genes in farm environments and food processing environments.

Chemical analysis and sensory evaluation of mahi-mahi (Cryphaena hippurus) during chilled storage.

Biogenic amines, total volatile base-nitrogen (TVB-N), and sensory evaluation are some of the indicators used for fish quality determination. Our objective was to evaluate the relationship among histamine, cadaverine, putrescine, TVB-N, and sensory evaluation as quality assessment tools. Two groups of six mahi-mahi fillets were refrigerated at 7 degrees C and sampled on days 0, 2, 4, 6, 8, and 10. On day 3, histamine, cadaverine, and putrescine levels reached 5, 3, and 0.5 mg/100 g, respectively, whereas TVB-N reached 30 mg/100 g. Sensory scores were 6 to 6.5 (10 very fresh and 1 very spoiled) for odor, appearance, texture, and color. Correlations were 0.78 and 0.72 between histamine and cadaverine and histamine and putrescine, 0.74 and 0.80 between TVB-N and cadaverine and TVB-N and putrescine, and 0.75 and 0.78 between odor and putrescine and odor and cadaverine. AromaMaps showed distinct trends for deteriorating mahi-mahi (Coryphaena hippurus) quality.

Identification of bacteria crucial to histamine accumulation in pacific mackerel during storage.

Bacterial growth and histamine formation in Pacific mackerel during storage at 0, 4, 15, and 25 degrees C were monitored. To identify bacterial species contributing to histamine formation, several groups of bacteria were isolated by using selective media under temperatures corresponding to the various storage conditions. Initially, low counts of bacteria were found in the gill, skin, and intestine of fresh fish, and only weak histamine formers were found in the gill. Histamine was found in the muscle when fish were stored above 4 degrees C, and aerobic plate counts reached 10(6) CFU/g. When fish became unsuitable for human consumption by abusive storage, toxicological levels of histamine were always found. The highest level of histamine formed was 283 mg/100 g in 2 days. The optimum temperature for supporting growth of prolific histamine formers was 25 degrees C. The most prolific and prevalent histamine former was Morganella morganii, followed by Proteus vulgaris, both of which were isolated on violet red bile glucose (VRBG) agar. At 15 degrees C, a significant level of histamine was still produced in fish muscle, although prolific histamine formers were less frequently detected than at 25 degrees C. The isolates on thiosulfate citrate bile salts sucrose (TCBS) agar were weak histamine formers and identified as Vibrio parahaemolyticus and Vibrio alginolyticus. At 4 degrees C, less than 57.4 mg/100 g of histamine was found in fish stored for 14 days. Most isolates were natural bacterial flora in the marine environment and identified as weak histamine formers. At 0 degrees C, neither histamine former nor histamine production was detected up to 14 days of storage.

Destruction of Escherichia coli O157:H7 and Salmonella enteritidis in cow manure composting.

Application of cow manure and composted manure in agricultural practice could potentially cause contamination of foodstuffs with pathogenic bacteria such as Salmonella Enteritidis and Escherichia coli O157:H7. In this study, rifampicin-resistant (RifR) E. coli O157:H7 and Salmonella Enteritidis at a level of 7 log CFU/g of raw compost feed were used to determine the effect of a bench-scale composting system on their survival. RifR E. coli O157:H7 was not detected after 72 h of composting at 45 degrees C, and RifR Salmonella Enteritidis was not detected after 48 h. The use of selective media for enrichment failed to recover in the composting samples held at 45 degrees C for 96 h. However, the pathogens showed no change in bacterial numbers when the composting system was held at room temperature. Thus, properly composted manure can be safely used in food crop production while minimizing the likelihood of microbial contamination.

Source and identification of histamine-producing bacteria from fresh and temperature-abused albacore.

Histamine-producing bacteria were isolated from fresh and temperature-abused albacore using two different isolation procedures. Typically, the bacterial isolates on Niven's or modified Niven's medium produced negligible or low levels of histamine (<300 ppm) in histamine enumeration broth. The most frequently found species using this approach was Hafnia alvei. By prescreening on selective media (eosin methylene blue [EMB] agar for enteric bacteria; deMan Rogosa Sharpe agar for lactic acid bacteria: KF streptococcus agar for streptococci; pseudomonas isolation [PI] agar for pseudomonads; and staphylococcus medium 110 agar for staphylococci) prior to plating on histidine decarboxylase differential media, detection rate of true histamine formers increased. Prolific histamine producers capable of forming >1,000 ppm histamine in culture broth were isolated when PI and EMB agars were used for prescreening. Among the selective media tested, EMB agar was most effective in selecting high histamine producers, as demonstrated by the highest rate of true positives based on histamine analysis. Histamine-producing isolates were mostly enteric bacteria, including Morganella morganii, H. alvei, Klebsiella spp., Citrobacter freundii, Enterobacter spp., and Serratia spp. M. morganii isolated on PI agar from temperature-abused albacore muscle was found to be the highest histamine former. This species was not isolated from fresh albacore. while other enteric bacteria were frequently detected on the gills. However, only a few species isolated from both fresh and temperature-abused muscles were identified as high histamine formers.

Microbiological, sensory, and electronic nose evaluation of yellowfin tuna under various storage conditions.

Microbiological assessment, sensory evaluation, and electronic nose (AromaScan) analysis were performed on yellowfin tuna stored at 0, 4, 10, and 22 degrees C for 0, 1, 3, 5, and 9 days. Fish color, texture, appearance, and odor were evaluated by a trained sensory panel, while aroma-odor properties were evaluated using an AromaScan. Bacterial enumeration was performed using plate count agar containing 1.5% NaCl. Tuna fillets stored at 22 degrees C for 3 days or longer had a bacterial load of over 10(7) CFU/g and were rated not acceptable for consumption (grade C) by the sensory panel. Tuna fillets stored at 4 degrees C for 9 days or 10 degrees C for over 5 days were rated as grade C products and also had a bacterial load of over 10(7) CFU/g. The change in fish quality as determined by AromaScan followed increases in microbiological counts in tuna fillets, indicating that bacterial load can serve as a useful and objective indicator of gross spoilage. Electronic nose devices can be used in conjunction with microbial counts and sensory panels to evaluate the degree of decomposition in tuna during storage.

Mutagenicity studies of benzidine and its analogs: structure-activity relationships.

The Ames Salmonella/microsome assay was employed to test the mutagenicity of benzidine and its analogs using strains TA98 and TA100 in the presence and absence of Aroclor 1254-induced rat S9 mix. 3,3'-Dichlorobenzidine-2HCl and 4,4'-dinitro-2-biphenylamine were directly mutagenic to TA98, while 4,4'-dinitro-2-biphenylamine was directly mutagenic to both TA98 and TA100 in the absence of S9 mix. 2-Aminobiphenyl, 3-aminobiphenyl, and 3,3'-5,5'-tetramethylbenzidine were not mutagenic in either strains in the presence or absence of S9. In the presence of S9 mix, 4-aminobiphenyl, benzidine, 3, 3'-dichlorobenzidine-2HCl, 3,3'-dimethoxybenzidine, 3,3'-4, 4'-tetraaminobiphenyl, o-tolidine, N, N-N', N'-tetramethylbenzidine, and 4,4'-dinitro-2-biphenylamine were mutagenic to TA98; 4-aminobiphenyl, 3,3'-dichlorobenzidine-2HCl, 3, 3'-dimethoxybenzidine, and 4,4'-dinitro-2-biphenylamine were mutagenic to TA100. Physicochemical parameters of these compounds including oxidation potentials, the energy difference between the lowest unoccupied molecular orbital and the highest occupied molecular orbital, ionization potentials, dipole moment, relative partition coefficient, and basicity did not correlate with their bacterial mutagenic activities.

Antibacterial mechanism of allyl isothiocyanate.

Allyl isothiocyanate (AITC), a natural compound in plants belonging to the family Cruciferae, has been shown to have strong antimicrobial activity in liquid media as well as in its vapor form. To understand its antimicrobial mechanism, AITC was tested for bactericidal activities to Salmonella Montevideo, Escherichia coli O157:H7, and Listeria monocytogenes Scott A at different stages of growth and was compared with streptomycin, penicillin G, and polymyxin B, each of known antibacterial mechanisms. Bactericidal activities were determined by measuring bacterial viability and leakage of metabolites. To determine its effects on membrane permeability, beta-galactosidase activity was examined after exposure of E. coli K-12 strain 3.300 to the three antibiotics and to AITC. The two gram-negative bacteria, Salmonella Montevideo and E. coli O157:H7, were more sensitive to AITC and to polymyxin B than the gram-positive L. monocytogenes. AITC and polymyxin B were effective bactericidal agents to bacteria at all growth stages, whereas penicillin G and streptomycin did not exhibit bactericidal activity to stationary cells. High A260 and A280 values of cellular filtrate and beta-galactosidase activity were obtained after treatments of AITC and polymyxin B. These data indicated that AITC was most similar to polymyxin B with respect to its antibacterial effect on cell membranes and on leakage of cellular metabolites. Gaseous AITC caused metabolite leakages, measurable increases in 3-galactosidase activity, and reduction of viable bacteria. The effectiveness of AITC in inhibiting bacteria at all growth stages and its strong activity in vapor phase support its application in food preservation.

Bactericidal activity of isothiocyanate against pathogens on fresh produce.

The bactericidal activity of allyl and methyl isothiocyanate (AITC and MITC) was tested with a rifampicin-resistant strain of Salmonella Montevideo and streptomycin-resistant strains of Escherichia coil O157:H7 and Listeria monocytogenes Scott A. Iceberg lettuce inoculated with high (10(7) to 10(8) CFU/g) and low (10(3) to 10(4) CFU/g) concentrations of bacterial pathogens was treated with AITC and MITC in sealed containers at 4 degrees C for 4 days. AITC showed stronger bactericidal activity than MITC against E. coli O157:H7 and Salmonella Montevideo, whereas MITC showed stronger activity against L. monocytogenes than E. coli O157:H7 and Salmonella Montevideo. Up to 8-log reduction occurred with E. coli O157:H7 and Salmonella Montevideo on lettuce following treatment with vapor generated from 400 microl of AITC for 2 and 4 days, respectively. AITC was used to treat tomatoes inoculated with Salmonella Montevideo on stem scars and skin and apples inoculated with E. coli O157:H7 on stem scars. The bactericidal effect of AITC varied with bacteria species and exposure time. Salmonella Montevideo inoculated on tomato skin was more sensitive to AITC than that on stem scars. Treatment with vapor generated from 500 microl of AITC caused an 8-log reduction in bacteria on tomato skin but only a 5-log reduction on tomato stem scars. The bactericidal activity of AITC was weaker for E. coli O157:H7 on apple stem scars; only a 3-log reduction in bacteria occurred when 600 microl of AITC was used.

Retention and changes of soy isoflavones and carotenoids in immature soybean seeds (Edamame) during processing.

Isoflavones and carotenoids in four experimental genotypes and Hutcheson cultivar soybeans were evaluated as a function of processing treatments and maturity. Total isoflavone and carotenoid contents were affected by genotypes and maturity stages (p < 0.0001). Total isoflavones ranged from 472 microg/g (in NTCPR93-40) to 2280 microg/g (in Hutcheson). Lutein contents ranged from 895 (in NTCPR93-286) to 2119 (in Honey Brown), and beta-carotene ranged from 291 (in Hutcheson) to 491 (in NICPR92-40) microg/100 g. Mean total isoflavone retention percentages in immature Hutcheson soybeans were 46% (boiling), 53% (freezing), and 40% (freeze-drying). Mean retentions of lutein and beta-carotene, respectively, were 92 and 73% in frozen, 62 and 62% in boiled, and 34 and 27% in freeze-dried soybeans. Boiling caused a substantial increase in daidzin, genistin, and genistein. The results show that post-harvest changes in total isoflavones and carotenoids in soybeans are influenced by processing methods, but genotype has an effect on isoflavone and carotenoid profiles during seed development.

HPLC method for analysis of free amino acids in fish using o-phthaldialdehyde precolumn derivatization.

Precolumn derivatization applying o-phthaldialdehyde (OPA) was used to analyze free lysine, histidine, and ornithine, precursors of the respective biogenic amines cadaverine, histamine, and putrescine, which are considered indicators of fish quality and safety. This method uses 75% methanol to eliminate the use of strong acids as the extraction solution. Each analysis took 35 min, was reproducible, and allowed separation of primary amino acids in fish samples. A binary solvent delivery system coupled with a fluorescence detector and an Ultrasphere ODS column were utilized for HPLC separation. Linearity of the calibration curves was very good (r(2) = 0.99) for the amino acids of interest. Minimum concentrations of detection were 40 pmol/mL for histidine and lysine and 70 pmol/mL for ornithine. Average recoveries were 72% for lysine, 93% for histidine, and 98% for ornithine. This method used solvent gradient elution to study the levels of these analytes in mahi-mahi, bigeye tuna, and flounder.

Determination of chlorate and chlorite and mutagenicity of seafood treated with aqueous chlorine dioxide.

The use of chlorine dioxide (ClO(2)) as a potential substitute for aqueous chlorine to improve the quality of seafood products has not been approved by regulatory agencies due to health concerns related to the production of chlorite (ClO(2)(-)) and chlorate (ClO(3)(-)) as well as possible mutagenic/carcinogenic reaction products. Cubes of Atlantic salmon (Salmo salar) and red grouper (Epinephelus morio) were treated with 20 or 200 ppm aqueous chlorine or ClO(2) solutions for 5 min, and extracts of the treated fish cubes and test solutions were checked for mutagenicity using the Ames Salmonella/microsome assay. No mutagenic activity was detected in the treated fish samples or test solutions with ClO(2). Only the sample treated with 200 ppm chlorine showed weak mutagenic activity toward S. typhimurium TA 100. No chlorite residue was detected in sea scallops, mahi-mahi, or shrimp treated with ClO(2) at 3.9-34.9 ppm. However, low levels of chlorate residues were detected in some of the treated samples. In most cases, the increase in chlorate in treated seafood was time- and dose-related.

Tannins and human health: a review.

Tannins (commonly referred to as tannic acid) are water-soluble polyphenols that are present in many plant foods. They have been reported to be responsible for decreases in feed intake, growth rate, feed efficiency, net metabolizable energy, and protein digestibility in experimental animals. Therefore, foods rich in tannins are considered to be of low nutritional value. However, recent findings indicate that the major effect of tannins was not due to their inhibition on food consumption or digestion but rather the decreased efficiency in converting the absorbed nutrients to new body substances. Incidences of certain cancers, such as esophageal cancer, have been reported to be related to consumption of tannins-rich foods such as betel nuts and herbal teas, suggesting that tannins might be carcinogenic. However, other reports indicated that the carcinogenic activity of tannins might be related to components associated with tannins rather than tannins themselves. Interestingly, many reports indicated negative association between tea consumption and incidences of cancers. Tea polyphenols and many tannin components were suggested to be anticarcinogenic. Many tannin molecules have also been shown to reduce the mutagenic activity of a number of mutagens. Many carcinogens and/or mutagens produce oxygen-free radicals for interaction with cellular macromolecules. The anticarcinogenic and antimutagenic potentials of tannins may be related to their antioxidative property, which is important in protecting cellular oxidative damage, including lipid peroxidation. The generation of superoxide radicals was reported to be inhibited by tannins and related compounds. The antimicrobial activities of tannins are well documented. The growth of many fungi, yeasts, bacteria, and viruses was inhibited by tannins. We have also found that tannic acid and propyl gallate, but not gallic acid, were inhibitory to foodborne bacteria, aquatic bacteria, and off-flavor-producing microorganisms. Their antimicrobial properties seemed to be associated with the hydrolysis of ester linkage between gallic acid and polyols hydrolyzed after ripening of many edible fruits. Tannins in these fruits thus serve as a natural defense mechanism against microbial infections. The antimicrobial property of tannic acid can also be used in food processing to increase the shelf-life of certain foods, such as catfish fillets. Tannins have also been reported to exert other physiological effects, such as to accelerate blood clotting, reduce blood pressure, decrease the serum lipid level, produce liver necrosis, and modulate immunoresponses. The dosage and kind of tannins are critical to these effects. The aim of this review is to summarize and analyze the vast and sometimes conflicting literature on tannins and to provide as accurately as possible the needed information for assessment of the overall effects of tannins on human health.

Enhanced broth media for selective growth of Vibrio vulnificus.

Rapid detection of Vibrio vulnificus can be enhanced by optimizing the components of enrichment broth. PNC (5% peptone, 1% NaCl, and 0.08% cellobiose [pH 8.0]) enhanced the growth of V. vulnificus compared to alkaline peptone broth. PNCC (PNC with 1.0 to 4.1 U of colistin methanesulfonate per ml) increased the growth of low levels of V. vulnificus while suppressing non-target bacteria.

Effects of the nitro-group on the mutagenicity and toxicity of some benzamines.

The Ames Salmonella/microsomal assay was employed to test the mutagenicity of some benzamines (aniline, and o- and p-phenylenediamine) and their nitro-derivatives (p-nitroaniline, 2-nitro-p-phenylenediamine, 3- and 4-nitro-o-phenylenediamine), using strains TA98 and TA100 and their nitroreductase-deficient mutants, TA98NR and TA100NR, in the presence and absence of rat S9 mix. The addition of the nitro-group to benzamine molecules converted them into direct mutagens. Furthermore, the position of the nitro-group affected their mutagenic activities. Cytotoxicity testing with Chinese hamster ovary cells (CHO-K1) showed that the presence of the nitro-group in these compounds had no specific effect on toxicity. The test compounds all showed a dose-related increase in inducing chromosomal aberrations in CHO cells. However, the presence of the nitro-group did not affect potency in inducing chromosomal aberrations. Compounds containing the nitro-group had higher initial oxidation potentials and dipole moments (mu) than their nonnitro-containing counterparts. The mutagenicity and toxicity of these compounds were not related to physico-chemical properties, including oxidation potential, energy difference (deltaE) between the lowest unoccupied molecular orbital (LUMO) and the highest occupied molecular orbital (HOMO), ionization potential (I.P.), and mu.

Mutagenicity and toxicity studies of p-phenylenediamine and its derivatives.

The mutagenicity of p-phenylenediamine and its derivatives was tested using Ames Salmonella strains TA98 and TA100. p-Phenylenediamine was weakly mutagenic to TA98 with metabolic activation. 2-Nitro-p-phenylenediamine was directly mutagenic to both strains, while 2-methyl-p-phenylenediamine required S9 mix. All the test compounds induced a dose-related increase in chromosomal aberrations in Chinese hamster ovary (CHO) cells in the absence of the S9 mix. The mutagenicity and toxicity of these compounds did not correlate with their oxidation potentials, or any other tested physicochemical properties including the energy difference between the lowest unoccupied and the highest occupied molecular orbital, ionization potential, and dipole moment.

Growth and Survival of Salmonella montevideo on Tomatoes and Disinfection with Chlorinated Water.

The survival on tomato fruits ( Lycopersicum esculentum ) of a rifampicin-resistant strain of Salmonella montevideo (Centers for Disease Control and Prevention [CDC] isolate G4639), the alleged source of the 1993 multistate outbreak of salmonellosis, was affected by inoculum dose and inoculation site (unbroken surface or wounds and stem scars), as well as by the medium (distilled water, Butterfield's buffer, or trypticase soy broth [TSB]) used to deliver the bacterium, This bacterium inoculated at 4 log10 CFU (colony forming units) per site in distilled water survived for 20 h on tomato skin. However, comparable survival occurred at the stem scars and growth cracks with smaller inoculum doses (3 log10 CFU). The bacterial populations increased rapidly on puncture wounds and tomato slices but decreased on the unbroken surface and stem scar. With unbroken skin and approximately 4 log10 CFU per site, the population survived for at least 48 h but could not be consistently detected after 5 days. By contrast, the stem scar population survived for at least 7 days and decreased only 1 to 2 log10 units. The inherently low pH of the tomatoes did not inhibit bacterial growth. Treatment with 100 ppm of aqueous chlorine for up to 2 min failed to kill all bacteria at these inoculation sites. This was especially true when the bacterial suspensions were prepared in TSB. TSB supported better bacterial survival and/or growth and also protected against the bactericidal effect of aqueous chlorine.

Mutagenicity studies of kojic acid.

Kojic acid, a fungal metabolite produced by some species of Aspergillus and Penicillium, was found to induce sister chromatid exchanges and chromosomal aberrations in Chinese hamster ovary cells in the presence or absence of the rat liver S9 mix. Furthermore, this compound was demonstrated to induce mutations in Salmonella typhimurium strains TA98 and TA100 using both plate-incorporation and preincubation methods.