Carcinogenesls vol.13 no.8 pp.1437-1445, 1992
Response of the ke test to NCI/NTP-screened chemicals, n . Genotoxic carcinogens and non-genotoxic non-carcinogens
George Bakale and Richard D.McCreary Case Western Reserve University, Department of Radiology, Biochemical Oncology Division, Cleveland, OH 44106-5000, USA
Introduction The a priori identification of potential carcinogens among the myriad of chemicals to which we are daily exposed continues to pose a formidable problem that has challenged several •Abbreviations: STT, short-term test; NTP, National Toxicology Program; NCI, National Cancer Institute; [S], solute concentration; CMP, 3-chloro-2-methylpropene; AEA, 3-amino-4-ethoxyacetanilide; CASRN, Chemical Abstracts Service Registry Number; S/A, structural alert of electrophilicity; LE, level of carcinogenic effect; MTD, maximum tolerated dose; SAR, structure-activity relationship; M-L, mouse-liver.
© Oxford University Press
1437
Downloaded from http://carcin.oxfordjournals.org/ at Cambridge University on August 15, 2015
A physico-chemical carcinogen-screening test was used to measure the rate constants of electron attachment, Jt^s, of 105 chemicals that had been screened in long-term rodent bioassays and short-term in vitro tests by the NCI/NTP. In the &e test, a pulse-conductivity technique is used to generate and monitor the decay of excess electrons that serve as nucleophilk surrogates for the target tissue of rodents. Of the 61 chemicals that had been found to be rodent carcinogens as well as Salmonella mutagens, 36 yield k,s that are equal to or greater than the diffusion-controlled kt of carbon tetrachloride and are considered to be positive kt test responses. In contrast, 29 of the remaining 44 chemicals that are putative non-carcinogens and non-mutagens yield &eS that are negative k? test responses. These results are combined with the ke responses of 46 non-mutagenic carcinogens and 20 mutagenic non-carcinogens that were reported earlier and are evaluated to determine the degree to which the measure of electron-accepting capacity that &,, provides complements or overlaps the electrophilkity or DNA reactivity of chemicals that is indicated by positive mutagenkity responses in the Ames Salmonella tester strains or by positive structural alerts, S/As, of the chemicals. The combined t, test results indicate that the overall predktivity of the kt test is comparable to and complements the Ames Salmonella test and S/As in identifying rodent carcinogens. Moreover, the electrons serve as non-discriminate nuckophilic targets for both genotoxic and non-genotoxk electronaccepting molecules and appear to attach with equal efficiency to carcinogens that are active in various tissues of rodents. This property of excess electrons suggests that the predictivity of the kc test could be enhanced by combining the measured kt with an appropriate lipophilkity or pharmacokinetic parameter. A pre-chemical electron-transfer step that had been proposed to precede chemical interactions between the carcinogen and target tissue is discussed in light of recent developments in electron-donor/-acceptor chemistry and in the application of structure—activity relationships to identify carcinogens.
disciplines for decades. Responses to this challenge have ranged from the quantum chemical studies of the Pullmans (1) to the biochemical studies of the metabolic activation of carcinogens by the Millers (2), and during the past 15 years the response of genetic toxicologists has focused on attempts to develop shortterm tests (STTs*) that would emulate or complement the most dominant STT, die Ames Salmonella microsome assay (3). The discipline employed in this work is most appropriately labeled liquid-state electronics, a field in which the unique transport and reaction properties of excess electrons in non-polar liquids are exploited to obtain a variety of information (4). The primary objective of the current study and our previous studies of potential carcinogens and putative non-carcinogens (5—8) was to determine if k^, the electron attachment rate constant, provides a measure of the electron-accepting properties of these chemicals that is sufficiently concordant with chemical carcinogenicity to be of value in identifying potential carcinogens. A secondary objective was to determine the degree to which k^ responses complement mutagenicity responses of the Ames Salmonella test. The rationale of using k^ as an indicator of chemical carcinogenicity was noted in our earlier studies (5—9) and was described in greater detail in two reviews (10,11). This rationale can be most simply summarized by regarding excess electrons in die ^ t e s t ^ a surrogate for the electron-rich target tissue of rodents in bioassays or for the DNA 'hot spots' of Salmonella tester strains in the Ames test. Whereas the nucleophilic biotargets of die initiation of carcinogenesis in animals and of mutagenesis in bacteria are not sufficiently nucleophilic to interact with procarcinogens that have not been metabolically activated, highly mobile and reactive excess electrons attach at every encounter to many unactivated procarcinogens. Thus, the diffusioncontrolled attachment of" the most fundamental nucleophile, an electron, to a test chemical indicates a high probability of that chemical being sufficiently electrophilic to be a potential carcinogen. Quantitation of this probability was another objective of measuring the k,s of chemicals that had been screened via the National Toxicology Program (NTP) in this and our previous study (9), which is hereafter denoted as I. In study I, k^s were measured in diose chemicals that had been reported by Zeiger to be non-mutagenic carcinogens or mutagenic non-carcinogens in a survey of 224 chemicals (12). These two-year, two-species rodent bioassays were conducted under die aegis of the National Cancer Institute (NCI) and the NTP who also sponsored mutagenicity screening of the same chemicals with the Ames Salmonella test {13). The NCI/NTP studies provide a unique database that has been used to draw significant conclusions regarding relationships among bacterial mutagenicity, rodent carcinogenicity and chemical structure (e.g. 13,14). The k^ results reported herein are entirely for genotoxic carcinogens and non-genotoxic non-carcinogens which balance the k^ results reported in I. Combining the two k,. studies provides novel physico-chemical information on the interaction of electrons with both types of carcinogens and non-carcinogens which should complement the aforementioned
G.Bakale and R.D.McCreary
carcinogenicity/mutagenicity/chemical-structure studies. The NTP-screened chemicals that were reported by Zeiger to yield equivocal carcinogenicity responses in rodents will be the subject of a forthcoming paper.
Table II. Electron attachment rate constants, k^, structural alerts of carcinogenic activity, S/As and levels of carcinogenic effect, LEs, of NCI/NTP-tested rodent carcinogens reported by Zeiger (12) to yield positive responses in the Ames Salmonella test Chemical (source)*
Materials and methods The pulse-conductivity system, solvent-purification technique and samplepreparation protocols that were used in I and had been described in detail in earlier studies (7,11) were again used to measurefcjSin this work. Excess electrons were produced in the cyclohexane solution of test chemical at 21 ± 2°C by a single 15 ns pulse of 1 MeV electrons from a Van de Graaff generator, and the half-life, r w , of electrons attaching to the test solute was directly measured in the 70—700 ns time regime using an oscilloscope. From the observed t^ and the solute concentration [S], Jtc was obtained via = In
(1)
Table I. Chemicals reported as mutagenic carcinogens or non-mutagenic non-carcinogens by Zeiger (12) that could not be screened with the k^ test Chemical (source*)
CASRN15
Comment0
(A) Mutagenic carcinogens Basic Red 9HC1(S) 3-Chloro-2-methylpropene (TK)d D & C Red 9 (Ra) 2,4-DiaminoanisoleSO4 (Ra) Diglycidyl resorcinol ether Direct Black (CS) Direct Brown 95 (Ra) 1,2-Propylene oxide Ziram (Ra)
569-61-9 563-17-3 5160-02-1 39156-41-7 101-90-6 1937-37-7 16071-86-6 75-56-9 137-30-4
insoluble volatile insoluble insoluble unavailable insoluble insoluble volatile insoluble
(B) Non-mutagenic non-carcinogens EDTA 3Na (Fl) Pigment Yellow (Ra) Sodium diethyldithiocarbonate (CS) Titanium dioxide (PB) Witch hazel (Ra)
150-38-9 6358-85-6 148-18-5 13463-67-7 68916-39-2
insoluble insoluble insoluble insoluble complex mixture'
"Sources of chemicals for which ^ screening was attempted are: CS, Chem Service; Fl, Fluka; PB, Pfaltz and Bauer; Ra, Radian; Si, Sigma; TK, Tokyo Kasei. b Chemical Abstracts Service Registry Number. TJetails of solubility, volatility and availability provided in text. 'Classification of mutagenic response as 'equivocal' by Zeiger (12) revised to 'positive' by Ashby et al. (15). e Complex mixture having unspecified concentrations of active components.
1438
*, < 3.0 x 1012 3-Amino-4-ethoxyacetanilide (Ra) 3-Amino-9-ethylcarbazole (Si) o-Anisidine (CS) Azobenzene (CS) Bis(2-chloro-l-methylethyl)ether (Ra) 2-ChloromethylpyridineHCl (PB) 4-Chloro-m-phenylenediamine (PB) 4-Chloro-ophenylenediamine (CS) Cytembena (Ra) p-Cresidine (PB) 2,4-Diaminotoluene (PB) Dimethyl hydrogenphosphite (PB) Direct Blue 6 (PB) Disperse Blue 1 (Al) HC Blue 1 (Ra) Hydrazobenzene (Al) 4,4'-Methylenedianiline-2HCl (CS) 6-Nitrobenzimidazole (PB/TK) 4,4'-Oxydianiline (Si) Phenoxybenzamine HO (TK) Pivalolactone (Ra) 4,4'-Thiodianiline (Al) o-Toluidine (CS)
117-79-3 9041-5 82-28-0 119-34-6 121-664 12448-1 135-20-6 96-12-8 106-934 107-06-2 609-20-1 78-87-5 542-75-6 62-73-7 91-93-0 121-14-2 283240-8 303-344 129-15-7 90-94-8 2243-62-1 139-94-6 602-87-9 1777-84-0 99-59-2 1836-75-5 5307-14-2 156-10-5 99-55-8 105-11-3 842-07-9 95-06 584-84-9 8001-35-2 1582-09-8 126-72-7
3.9 3.8 4.2 4.2 3.3 3.2 3.3 3.7 3.6 5.1 3.4 3.5 3.4 3.3 4.4 4.7 3.3 3.1 4.2 4.6 3.3 32 5.2 3.4 5.5 5.6 3.5 5.3 6.0 3.1 4.1 5.0 5.5 4.8 3.6 4.1
+ + + + + +« + + + + + +8 + + + + + -h + + + + + + + + + + + + +8 + +8 + + +
A D A D D D8 A A A A C C8 A A B B A B1 A A A A A D A A D A B D C« A A8
17026-81-2 6109-97-3 90-04-0 103-33-3 108-60-1 6959484 5131-60-2 95-83-0 21739-91-3 120-71-8 95-80-7 868-85-9 260246-2 247545-8 2784-94-3 122-66-7 1355244-8 94-52-0 101-804 63-92-3 195545-9 139-65-1 95-534
0.7 2.5 0.1 1.6 1.0 1.9 1.0 0.9 0.1 0.1 0.2 0.1 0.4 0.6 1.9 0.6 0.1 2.3 0.2 1.1 0.2 0.5 0.03
+ + + + + + + + -h + + + +h + + + + + + +h + + +
D A A B B A A A B1 A A B BT C A A A C A A1 C A A
C B A
Downloaded from http://carcin.oxfordjournals.org/ at Cambridge University on August 15, 2015
A 2 nM concentration of an efficient electron acceptor such as the carcinogen carbon tetrachloride, CC14, is sufficient to reduce t^ to —100 ns from the 700 ns value of (w that is routinely observed in purified cyclohexane (Fisher Scientific, 99 mol%). As in study I, several solvents were used to dissolve test chemicals that were not sufficiently soluble in cyclohexane; these secondary solvents included ethanol, ethyl acetate, benzene, tetrahydrofuran and dimethyl formamide. Chemicals that were not soluble in these solvents at the required millimolar concentration prior to being diluted to the micromolar range in purified cyclohexane or which precipitated upon dilution are listed in Table I. Also in Table I are two chemicals, 3-chloro-2-methylpropene (CMP) and 1,2-propylene oxide, that are too volatile to permit k^s to be measured using the argon-bubbling degassing technique that we use. The 102 torr vapor pressure of CMP at ambient temperature also contributed to the 'equivocal' mutagenic response that Zeiger reported (12), which was revised to positive by Ashby et al. (15) based on a study by Warner et al. who obtained a positive mutagenic response when CMP was tested using a desiccator technique (16). The 'unavailable' chemical listed in Table I, diglycidyl resorcinol ether, could not be obtained from commercial sources nor from Radian Corporation, the repository of chemicals screened by the NTP. The unknown composition of witch hazel precluded measurement of the kc of this complex mixture.
*, >3.0 x 1012 M " 1 s" 1 2-Aminoanthraquinone (TK) 2-Aminobiphenyl (Al) l-Amino-2-methylanthraquinone (Ra) 4-Amino-2-nitrophenol (Al) 2-Amino-5-nitrothiazole (Si) Chlorodibromomethane (Ra)f Cupferron (Ra) 1,2-Dibromo-3-chloropropane (CS) 1,2-Dibromomethane (Al) 1,2-Dichloroethane (CS) 2,6-Dichloro-p-phenylenediamine (Si) 1,2-Dichloropropane (CS) 1,3-Dichloropropene (CS) Dichlorvos (Ra)* Dimethoxybenzkline-4,4'-diisocyanate (PB) 2,4-Dinitrotoluene (CS) Disperse Yellow 3 (Si) Lasiocarpine (Ra) 2-Methyl-l-rutroanthraquinone (Ra) Michler's ketone (CS) 1,5-Naphthalenediamine (PB) Nithiazide (Ra) 5-Nitroacenaphthene (Al) 3-Nitro-/?-acetophenetide (Ra) 5-Nitro-o-anisidine (Si) Nitrofen (Ra) 2-Nitro-p-phenylenediamine (CS) p-Nitrosodiphenylamine (PB) 5-Nitro-o-toluidine (CS) p-Quinone dioxime (PB) Solvent Yellow 14 (Si) Sulfallate (CS) 2,4-Toluene diisocyanate (Si)1 Toxaphene (CS) Trifluralin (Ra) Tris(2,3-dibromopropylphosphate) (Al)
S/Ad
CASRNb
*. test on NCI/NTP-screened chemicals
Table n . continued 8
Chemical (source)
2,4,5-Trimethylanilinc (CS) 2,6-Xylidine (PB)k
CASRN*
kc
S/Ad LE"-e
137-17-7 0.01 + h 87-62-7 0.02 +
A B1
Results The k^ values of 61 rodent carcinogens that Zeiger reported to yield positive Ames test responses (12) are listed in Table II. As in study I, the chemical nomenclature used by Zeiger is adhered to in Table II, and the k^ are again divided into two categories: k
Response of the ke test to NCI/NTP-screened chemicals, n . Genotoxic carcinogens and non-genotoxic non-carcinogens
George Bakale and Richard D.McCreary Case Western Reserve University, Department of Radiology, Biochemical Oncology Division, Cleveland, OH 44106-5000, USA
Introduction The a priori identification of potential carcinogens among the myriad of chemicals to which we are daily exposed continues to pose a formidable problem that has challenged several •Abbreviations: STT, short-term test; NTP, National Toxicology Program; NCI, National Cancer Institute; [S], solute concentration; CMP, 3-chloro-2-methylpropene; AEA, 3-amino-4-ethoxyacetanilide; CASRN, Chemical Abstracts Service Registry Number; S/A, structural alert of electrophilicity; LE, level of carcinogenic effect; MTD, maximum tolerated dose; SAR, structure-activity relationship; M-L, mouse-liver.
© Oxford University Press
1437
Downloaded from http://carcin.oxfordjournals.org/ at Cambridge University on August 15, 2015
A physico-chemical carcinogen-screening test was used to measure the rate constants of electron attachment, Jt^s, of 105 chemicals that had been screened in long-term rodent bioassays and short-term in vitro tests by the NCI/NTP. In the &e test, a pulse-conductivity technique is used to generate and monitor the decay of excess electrons that serve as nucleophilk surrogates for the target tissue of rodents. Of the 61 chemicals that had been found to be rodent carcinogens as well as Salmonella mutagens, 36 yield k,s that are equal to or greater than the diffusion-controlled kt of carbon tetrachloride and are considered to be positive kt test responses. In contrast, 29 of the remaining 44 chemicals that are putative non-carcinogens and non-mutagens yield &eS that are negative k? test responses. These results are combined with the ke responses of 46 non-mutagenic carcinogens and 20 mutagenic non-carcinogens that were reported earlier and are evaluated to determine the degree to which the measure of electron-accepting capacity that &,, provides complements or overlaps the electrophilkity or DNA reactivity of chemicals that is indicated by positive mutagenkity responses in the Ames Salmonella tester strains or by positive structural alerts, S/As, of the chemicals. The combined t, test results indicate that the overall predktivity of the kt test is comparable to and complements the Ames Salmonella test and S/As in identifying rodent carcinogens. Moreover, the electrons serve as non-discriminate nuckophilic targets for both genotoxic and non-genotoxk electronaccepting molecules and appear to attach with equal efficiency to carcinogens that are active in various tissues of rodents. This property of excess electrons suggests that the predictivity of the kc test could be enhanced by combining the measured kt with an appropriate lipophilkity or pharmacokinetic parameter. A pre-chemical electron-transfer step that had been proposed to precede chemical interactions between the carcinogen and target tissue is discussed in light of recent developments in electron-donor/-acceptor chemistry and in the application of structure—activity relationships to identify carcinogens.
disciplines for decades. Responses to this challenge have ranged from the quantum chemical studies of the Pullmans (1) to the biochemical studies of the metabolic activation of carcinogens by the Millers (2), and during the past 15 years the response of genetic toxicologists has focused on attempts to develop shortterm tests (STTs*) that would emulate or complement the most dominant STT, die Ames Salmonella microsome assay (3). The discipline employed in this work is most appropriately labeled liquid-state electronics, a field in which the unique transport and reaction properties of excess electrons in non-polar liquids are exploited to obtain a variety of information (4). The primary objective of the current study and our previous studies of potential carcinogens and putative non-carcinogens (5—8) was to determine if k^, the electron attachment rate constant, provides a measure of the electron-accepting properties of these chemicals that is sufficiently concordant with chemical carcinogenicity to be of value in identifying potential carcinogens. A secondary objective was to determine the degree to which k^ responses complement mutagenicity responses of the Ames Salmonella test. The rationale of using k^ as an indicator of chemical carcinogenicity was noted in our earlier studies (5—9) and was described in greater detail in two reviews (10,11). This rationale can be most simply summarized by regarding excess electrons in die ^ t e s t ^ a surrogate for the electron-rich target tissue of rodents in bioassays or for the DNA 'hot spots' of Salmonella tester strains in the Ames test. Whereas the nucleophilic biotargets of die initiation of carcinogenesis in animals and of mutagenesis in bacteria are not sufficiently nucleophilic to interact with procarcinogens that have not been metabolically activated, highly mobile and reactive excess electrons attach at every encounter to many unactivated procarcinogens. Thus, the diffusioncontrolled attachment of" the most fundamental nucleophile, an electron, to a test chemical indicates a high probability of that chemical being sufficiently electrophilic to be a potential carcinogen. Quantitation of this probability was another objective of measuring the k,s of chemicals that had been screened via the National Toxicology Program (NTP) in this and our previous study (9), which is hereafter denoted as I. In study I, k^s were measured in diose chemicals that had been reported by Zeiger to be non-mutagenic carcinogens or mutagenic non-carcinogens in a survey of 224 chemicals (12). These two-year, two-species rodent bioassays were conducted under die aegis of the National Cancer Institute (NCI) and the NTP who also sponsored mutagenicity screening of the same chemicals with the Ames Salmonella test {13). The NCI/NTP studies provide a unique database that has been used to draw significant conclusions regarding relationships among bacterial mutagenicity, rodent carcinogenicity and chemical structure (e.g. 13,14). The k^ results reported herein are entirely for genotoxic carcinogens and non-genotoxic non-carcinogens which balance the k^ results reported in I. Combining the two k,. studies provides novel physico-chemical information on the interaction of electrons with both types of carcinogens and non-carcinogens which should complement the aforementioned
G.Bakale and R.D.McCreary
carcinogenicity/mutagenicity/chemical-structure studies. The NTP-screened chemicals that were reported by Zeiger to yield equivocal carcinogenicity responses in rodents will be the subject of a forthcoming paper.
Table II. Electron attachment rate constants, k^, structural alerts of carcinogenic activity, S/As and levels of carcinogenic effect, LEs, of NCI/NTP-tested rodent carcinogens reported by Zeiger (12) to yield positive responses in the Ames Salmonella test Chemical (source)*
Materials and methods The pulse-conductivity system, solvent-purification technique and samplepreparation protocols that were used in I and had been described in detail in earlier studies (7,11) were again used to measurefcjSin this work. Excess electrons were produced in the cyclohexane solution of test chemical at 21 ± 2°C by a single 15 ns pulse of 1 MeV electrons from a Van de Graaff generator, and the half-life, r w , of electrons attaching to the test solute was directly measured in the 70—700 ns time regime using an oscilloscope. From the observed t^ and the solute concentration [S], Jtc was obtained via = In
(1)
Table I. Chemicals reported as mutagenic carcinogens or non-mutagenic non-carcinogens by Zeiger (12) that could not be screened with the k^ test Chemical (source*)
CASRN15
Comment0
(A) Mutagenic carcinogens Basic Red 9HC1(S) 3-Chloro-2-methylpropene (TK)d D & C Red 9 (Ra) 2,4-DiaminoanisoleSO4 (Ra) Diglycidyl resorcinol ether Direct Black (CS) Direct Brown 95 (Ra) 1,2-Propylene oxide Ziram (Ra)
569-61-9 563-17-3 5160-02-1 39156-41-7 101-90-6 1937-37-7 16071-86-6 75-56-9 137-30-4
insoluble volatile insoluble insoluble unavailable insoluble insoluble volatile insoluble
(B) Non-mutagenic non-carcinogens EDTA 3Na (Fl) Pigment Yellow (Ra) Sodium diethyldithiocarbonate (CS) Titanium dioxide (PB) Witch hazel (Ra)
150-38-9 6358-85-6 148-18-5 13463-67-7 68916-39-2
insoluble insoluble insoluble insoluble complex mixture'
"Sources of chemicals for which ^ screening was attempted are: CS, Chem Service; Fl, Fluka; PB, Pfaltz and Bauer; Ra, Radian; Si, Sigma; TK, Tokyo Kasei. b Chemical Abstracts Service Registry Number. TJetails of solubility, volatility and availability provided in text. 'Classification of mutagenic response as 'equivocal' by Zeiger (12) revised to 'positive' by Ashby et al. (15). e Complex mixture having unspecified concentrations of active components.
1438
*, < 3.0 x 1012 3-Amino-4-ethoxyacetanilide (Ra) 3-Amino-9-ethylcarbazole (Si) o-Anisidine (CS) Azobenzene (CS) Bis(2-chloro-l-methylethyl)ether (Ra) 2-ChloromethylpyridineHCl (PB) 4-Chloro-m-phenylenediamine (PB) 4-Chloro-ophenylenediamine (CS) Cytembena (Ra) p-Cresidine (PB) 2,4-Diaminotoluene (PB) Dimethyl hydrogenphosphite (PB) Direct Blue 6 (PB) Disperse Blue 1 (Al) HC Blue 1 (Ra) Hydrazobenzene (Al) 4,4'-Methylenedianiline-2HCl (CS) 6-Nitrobenzimidazole (PB/TK) 4,4'-Oxydianiline (Si) Phenoxybenzamine HO (TK) Pivalolactone (Ra) 4,4'-Thiodianiline (Al) o-Toluidine (CS)
117-79-3 9041-5 82-28-0 119-34-6 121-664 12448-1 135-20-6 96-12-8 106-934 107-06-2 609-20-1 78-87-5 542-75-6 62-73-7 91-93-0 121-14-2 283240-8 303-344 129-15-7 90-94-8 2243-62-1 139-94-6 602-87-9 1777-84-0 99-59-2 1836-75-5 5307-14-2 156-10-5 99-55-8 105-11-3 842-07-9 95-06 584-84-9 8001-35-2 1582-09-8 126-72-7
3.9 3.8 4.2 4.2 3.3 3.2 3.3 3.7 3.6 5.1 3.4 3.5 3.4 3.3 4.4 4.7 3.3 3.1 4.2 4.6 3.3 32 5.2 3.4 5.5 5.6 3.5 5.3 6.0 3.1 4.1 5.0 5.5 4.8 3.6 4.1
+ + + + + +« + + + + + +8 + + + + + -h + + + + + + + + + + + + +8 + +8 + + +
A D A D D D8 A A A A C C8 A A B B A B1 A A A A A D A A D A B D C« A A8
17026-81-2 6109-97-3 90-04-0 103-33-3 108-60-1 6959484 5131-60-2 95-83-0 21739-91-3 120-71-8 95-80-7 868-85-9 260246-2 247545-8 2784-94-3 122-66-7 1355244-8 94-52-0 101-804 63-92-3 195545-9 139-65-1 95-534
0.7 2.5 0.1 1.6 1.0 1.9 1.0 0.9 0.1 0.1 0.2 0.1 0.4 0.6 1.9 0.6 0.1 2.3 0.2 1.1 0.2 0.5 0.03
+ + + + + + + + -h + + + +h + + + + + + +h + + +
D A A B B A A A B1 A A B BT C A A A C A A1 C A A
C B A
Downloaded from http://carcin.oxfordjournals.org/ at Cambridge University on August 15, 2015
A 2 nM concentration of an efficient electron acceptor such as the carcinogen carbon tetrachloride, CC14, is sufficient to reduce t^ to —100 ns from the 700 ns value of (w that is routinely observed in purified cyclohexane (Fisher Scientific, 99 mol%). As in study I, several solvents were used to dissolve test chemicals that were not sufficiently soluble in cyclohexane; these secondary solvents included ethanol, ethyl acetate, benzene, tetrahydrofuran and dimethyl formamide. Chemicals that were not soluble in these solvents at the required millimolar concentration prior to being diluted to the micromolar range in purified cyclohexane or which precipitated upon dilution are listed in Table I. Also in Table I are two chemicals, 3-chloro-2-methylpropene (CMP) and 1,2-propylene oxide, that are too volatile to permit k^s to be measured using the argon-bubbling degassing technique that we use. The 102 torr vapor pressure of CMP at ambient temperature also contributed to the 'equivocal' mutagenic response that Zeiger reported (12), which was revised to positive by Ashby et al. (15) based on a study by Warner et al. who obtained a positive mutagenic response when CMP was tested using a desiccator technique (16). The 'unavailable' chemical listed in Table I, diglycidyl resorcinol ether, could not be obtained from commercial sources nor from Radian Corporation, the repository of chemicals screened by the NTP. The unknown composition of witch hazel precluded measurement of the kc of this complex mixture.
*, >3.0 x 1012 M " 1 s" 1 2-Aminoanthraquinone (TK) 2-Aminobiphenyl (Al) l-Amino-2-methylanthraquinone (Ra) 4-Amino-2-nitrophenol (Al) 2-Amino-5-nitrothiazole (Si) Chlorodibromomethane (Ra)f Cupferron (Ra) 1,2-Dibromo-3-chloropropane (CS) 1,2-Dibromomethane (Al) 1,2-Dichloroethane (CS) 2,6-Dichloro-p-phenylenediamine (Si) 1,2-Dichloropropane (CS) 1,3-Dichloropropene (CS) Dichlorvos (Ra)* Dimethoxybenzkline-4,4'-diisocyanate (PB) 2,4-Dinitrotoluene (CS) Disperse Yellow 3 (Si) Lasiocarpine (Ra) 2-Methyl-l-rutroanthraquinone (Ra) Michler's ketone (CS) 1,5-Naphthalenediamine (PB) Nithiazide (Ra) 5-Nitroacenaphthene (Al) 3-Nitro-/?-acetophenetide (Ra) 5-Nitro-o-anisidine (Si) Nitrofen (Ra) 2-Nitro-p-phenylenediamine (CS) p-Nitrosodiphenylamine (PB) 5-Nitro-o-toluidine (CS) p-Quinone dioxime (PB) Solvent Yellow 14 (Si) Sulfallate (CS) 2,4-Toluene diisocyanate (Si)1 Toxaphene (CS) Trifluralin (Ra) Tris(2,3-dibromopropylphosphate) (Al)
S/Ad
CASRNb
*. test on NCI/NTP-screened chemicals
Table n . continued 8
Chemical (source)
2,4,5-Trimethylanilinc (CS) 2,6-Xylidine (PB)k
CASRN*
kc
S/Ad LE"-e
137-17-7 0.01 + h 87-62-7 0.02 +
A B1
Results The k^ values of 61 rodent carcinogens that Zeiger reported to yield positive Ames test responses (12) are listed in Table II. As in study I, the chemical nomenclature used by Zeiger is adhered to in Table II, and the k^ are again divided into two categories: k
