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A compilation of genotoxicity and carcinogenicity data on aromatic aminosulphonic acids
0278-6915/92 $5.00 + 0.00 Copyright © 1992 Pergamon Press Ltd
Fd Chem. Toxw Vol 30, No. 7, pp 635-660, 1992 Pnnted m Great Britain All rights reserved
Review Section A COMPILATION OF GENOTOXICITY AND CARCINOGENICITY DATA ON AROMATIC AMINOSULPHONIC ACIDS R. JUNG, D. STEINLE* and R. ANLIKER* Hoechst AG, Gewerbetoxlkologle, PO Box 800320, D-6230 Frankfurt am Main, Germany and *Ecological and Toxicological Assocmtlon of Dyes and Orgamc Pigments Manufacturers (ETAD), PO Box, CH-4005, Basel, Switzerland (Accepted 16 March 1992)
Summary--A review is presented to evaluate existing information on genotox~city and carcinogenicity testing of various aromatic aminosulphomc acids (AASAs) A great variety of water-soluble azo dyes can form aromatic phenyl- or naphthyl-ammosulphomc acids by ebemlcal and enzymatic reduction. AASAs are also used as intermediates in the synthes~sof azo dyes and azo p~gments and can anse as contaminants in the final products Comparisons have been made with the data available on the corresponding unsulphonated analogues, some of which are known to be genotox~c and/or carcinogemc The vast majority of the AASAs were concluswely non-mutagenic in the Ames test. In most cases the absence of genotoxic~tywas also demonstrated with a variety of other test systems m wtro and m vivo It is concluded that AASAs, m contrast with some of their unsulphonated analogues, generally have no or very low genotoxtc and tumorigemc potentml.
Introduction Epidemiological data have shown an association between occupational exposure to 2-naphthylamme (no. 118 in Tables), benzidme (115), 4-aminobiphenyl (114) and 2-methyl-4-chloroaniline, and the risk of cancer of the urinary bladder. The carcmogenicity effects in humans and animals of a number of lipophilic aromatic amines are well documented, and experimental evidence indicates that this carcinogenicity is dependent on metabolic actwation. On the other hand, several aromatic amines have been shown not to possess tumorigenic actwity even when they are positive in the Ames test, clearly indicating that carcinogenicity is not a common feature for all compounds in this class of chemicals. Aromatic amines are important intermediates m the synthesis of a great variety of products, such as plastic materials, pesticides, rubber chemicals, and colourings, primarily azo colourings. The production and use of the above-mentioned four carcinogenic amines has been abandoned by the major dyestuffs manufacturers, and in many industrialized countries their production and use are controlled by very stringent regulations. 2-Naphthylamine (118) also played an important role as a starting material in the synthesis of key intermediates for colourings, especially aminonaphthalenesulphonic acids. *To whom correspondence should be addressed. Abbreviation: AASA = aromatic armnosulphomc acid.
Alternative synthetic routes to these aminonaphthalenesulphonic acids were developed to circumvent the use of the hazardous 2-naphthylamine (Anliker, 1977). Undoubtedly, substituted aminonaphthalenesulphonic acids, and generally aromatic aminosulphonic acids (AASAs; often also called 'sulphonated aromatic amines') will continue to be major intermedmtes in the manufacture of organic colourings. The seeming structural similarity between AASAs and the unsulphonated analogues gave nse to the suspicion that they might also possess a similar tumorigemc activity. As in other classes of chemicals, some of the existing shortcomings of structure/activity analogies can also be shown with these groups of chemicals (Anliker, 1983). The sulphonic acid closely related to 2-naphthylamine (118), 2aminonaphthalene-1-sulphonic acid (22), did not produce any carcinogenic effects in a chronic feeding study in mice, even at high dose levels of 1000 mg/kg body weight (Della Porta, 1982). Theiss et al. (1981) stud~ed seven 1-amino- and 2-aminonaphthalenesulphonic acids in the lung adenoma test in the strain A mouse using lp administration. The authors concluded that neither l-naphthylamine (117) nor the four sulphonic acid derivatives of l-naphthylamine tested were tumorigenic. However, 2-naphthylamine and two out of the three sulphomc acid derivatives of 2-naphthylamine tested were believed to have produced a statistically significant tumour response. A later paper by the same group (Maronpot et al., 1986) 635
N~
I
88-21-1
C6HTNO3S
~
121--47-I
C6HvNO3S
SO3H
~ S 0 3 H
N~
121-57-3
C6H~NO3S
so3H
+
Structure
No.
Zetger (1988) NTP Rep (1986) Shimtzu (1983)
Zelger (1988) NTP Rep (1986) Chung (1981) Chung (1978) Vemtt (1976)
ETAD (1989) + (w) Zeiser (1988) + (w) NTP Rep (1986)
-
-
Ames lest data
Aromatic ammosulphonic acids
m
DNA
m - cyt
Yoshtml (1988)
Dienckx (1989)
Other data 101 C6HvN 62-53-3
No. -
test
Ames
Group 3
C+
m+ c+ m+ mmm+ m+ m c+
Other data
GMA haembmding cyt cyt DNA GMA GMA DNA haembmdiug
m - SAL m - SAL m - SAL
Unsulphonated analogues
Table I Mutagemctty and other toxtcity data on selected aromatic ammosulphomc actds and thetr unsulphonated analogues*
IARC (1987)
Fassma (1990) Btrner (1988) lshtdate (1988) Ishtdate (1988) Yoshimi (1988) Mitchell (1988) Wangenhelm (1988) yon der Hude (1988) Hamson (1987) Zetger (1987)
Shahm (1989) Ishtdate (1988) Zetger (1987)
Z
NH 2
88--44-8
CH,NOS
NH 2
~
137-51-9
C6H?NO6S2
SO3H
S03H
98-37-3
C6HTNO4S
H O 3 S / ~ O H
HO3S
+
Zeiger (1988) ETAD (1987) NTP Rep (1986)
ETAD (1988) Shtmlzu (1983)
m- DNA m - cyt m - DNA m- GMA
c- cell trans
Yoshlml (1988) ETAD (1988) ETAD (1988) ETAD (1986)
Styles(1973)
103 CeHTNO 95-55-6
102 CTHgN 106-49-0
+ (w)
+
c+ m +/-
haembmdm8 cyt
m SAL m + (w) SAL
DNA
[contd]
Hamson (1987) Kirchner (1982)
De Flora (1984) BIBRA (1984)
Birner (1988) Neumann (1988) Pai (1985) Garner (1984) Ce-~trone (1982) c+ c+ mcm+
haembmdmg haembmdmg GMA
NTP Rep (1984) ETAD (1981)
m ÷ (w) SAL m+ SAL
fj3
~s o
f~
o
No
88-23-3
C6H6NCIO4S
N~
5857-94-3
C6H6NC104S
HO
SO3H
6370-23-6
CI ~
CI
CH 3
CsHtlNO3S
HaC ~
NH 2
Structure
SO3H
NH 3
SO3H
Zelger (1988) NTP Rep (1986)
ETAD (1987)
Ames test data Chung (1981)
Aromatic ammosulphomc aods
Table
Other data
1--contd
106 C~H6CINO 95-85-2
105 C6H6CINO 38191-33-2
No 104 CsHHN 95-68-1
+ (w)
Other data SAL SAL SAL DNA haembmdmg DNA
m + (w) SAL m + (w) SAL
m+ cc-l+ Group 3
C+
m+ m+ m+ m+
Unsulphonated analogues Ames test +
Zelger (1988) NTP Rep (1986)
Zelger (1988) NTP Rep (1985a) Shlmlzu (1983) Wdhams (1989) Blrner (1988) Yoshlml (1988) Garner (1984) DETO (1980) IARC (1987)
z
oo
12
1!
10
SO3H
6471-78-9
CsHttNO4 S
NH 2
Cl"
88-51-7
C7HsCINO3S
CH 3
1
NH 2
~
88-53-9
CTHsCINO3 S
12I-t3
Cl ~ / N H a
CH30"
~
SO3H
SO3H
CH 3
ETAD (1986)
ETAD (1986) Shtmlzu (1985)
Chung (1981)
rn DNA m DNA
m- DNA m- cyt
YoshJml (1988) ETAD (1986)
Wflhams (1989) ETAD (1988)
109 C7HsCIN 95-74-9
108 CTHsC1N 7149-75-9
107 CaHItNO 120-71-8
+ (w)
pulm ad
SAL SAL
[contd]
Ennever (1986) NTP-TR-145 (1978) DNA
m+ c
NTP Rep (1989a) Zelger (1987) Ennever (1986)
IARC (1987)
Maronpot (1986) NTP Rep 85-002 (1985) Vaimo (1985) Garner (1984) NTP-TR- 142 (1978)
Zeiger (1988) NTP Rep (1986)
m + (w) SAL mSAL mSAL
Group 2B
C+
C+
C+
C+
c-
m+ m+
~D
> >
o
0
g~
O
15
14
13
No.
5123-63-7
CIoHIsNO3 S
32432-55-6
C9H|4N203S
CH 3
-~ NH 2
/
HO3S ~-
-,~
CH3
N%
3577-63-7
CTHTNOsS
SO3H
H3C ~
HOOC
NH 2
Structure
--
-
--
ETAD (1989)
ETAD (1989)
NTP Rep (1984)
Ames test data
AromaUc ammosulphomc actds Other data
Table l--contd
112 CIoH~sN 91-66-7
I11 CgH 14N2 3102-70-3
-
-
110 C7HTNO2 118-92-3
test
No
mmm-
Group 3
c-
m m m+ m+ m-
SAL SAL DNA
SAL SAL GMA cyt DNA
Other data
Unsulphonated analogues Ames
Zelger (1988) Shlmlzu (1983) Yoshlml (1988)
IARC (1987)
Zelger (1987) NTP Rep (1984b) Mitchell (1988) NTP Rep (1985b) Cesarone (1982) NTP-TR-36 (1978)
z o-
19
18
17
16
I
-
~
H~~
N
SO3H
~
I
117-61-3
CI2HI2N206S2
N
3365-90-0
CI2HI2N206S2
3365--89-7
CI2HuN03S
98-40-8
C~I13NO3S
SO3H
~
~
S%H
NHC2H5
-
ETAD (1989)
Ashby (1982)
ETAD(1989)
¢
cell trans Ashby (1978)
115
115 CI2HI2N 92-87-5
il4 CI2HIIN 92-67-1
113 CgHI3N 94-68-8
c + Group I
c + Group I
c + Group I
IARC (1987)
IARC (1987)
IARC (1987)
[contdl
>
O
)0
O X
o
22
21
81-16-3
CloH9NO3S
SO3H
84-86-6
CIoH9NO3S
SO3H
81-11-8
C14HI4N206S2
SO3H
Structure
2N-Q-c.=c.
No.
SO3H
NH 2
-
Le (1985) Pogodma (1984) Purchase (1978)
Kler (1986) Chung (1981)
NTP Rep (1990) ETAD (1989) Zelger (1987)
Ames test data
Aromatac amlnosulphomc acads
GMA cyt
c e-
m- DNA m - cyt
c
m m
ETAD (1989) ETAD (1989) Pogodma (1984) Della Porta (1982)
D E T O (1980)
ETAD (1989) NTP Rep (1989b)
Other data
Table 1--contd
118 CIoH9N 91-59-8
117 CIoH9N 134-32-7
116 C14HI4N2 621-96-5
No
+
+
test
Ames
c+
Group 3
m-
Group 1
DNA
GMA cyt cyt cyt
mm+ mm+ C-
SAL SAL SAL SAL
m+ m+ m+ m÷
Other data
Unsulphonated analogues
IARC (1987)
IARC (1987)
Fassma (1990) lsh~date (1988) Isbldate (1988) NTP Rep (1988) Vamlo (1985) Brambdla (1985)
lshldate (1988) Zelger (1988) NTP Rep (1986) Kler (1986)
z
bO
26
25
24
23
1 17-26-4
Cl0H9NO6S 2
SO3H
SOaH
86-60-2
Ct0H9NO3S
93-00-5
C)0H9NO3S
~
HO3S~
81-05-0
C1oHgNO3 S
SO3H
NH 2
NH2
-
-
-
-
-
-
Kmr (1986) Purchase (1978) Garner (1977)
Le (1985)
Le (1985)
Le (1985)
C-
c-
cell trans
c pulm ad
c pulm ad
Purchase (1978) Purchase (1978)
Thetss (1981)
Thelss (I981)
118
118
118
118
+
+
C+
C+
C+
C+
Group 1
Group 1
Group I
Group 1
[contd]
IARC (1987)
IARC (1987)
IARC (1987)
IARC (1987)
o
o
o
29
28
27
No.
HO3S I
118-03-6
CIoH9NO9S 3
v
SO3H
~
118-33-2
Cl0H9~O6S2
SO3H
86-65-7
C10HgNO6S 2
~
H03S ~
HO3S
SO3H
Structure
~SO3H
NH 2
NH2
-
-
-
ETAD (1989)
ETAD (1989)
ETAD (1989)
Ames test data
Aromatic ammosulphonlc acids
c +~ pulm ad Theiss (1981)
Other data
Table l---contd
I 18
118
118
No
+
+
Ames test
c+
c+
¢+
Group I
Group 1
Group 1
Other data
Unsulphonated analogues
IARC(1987)
IARC (1987)
IARC (1987)
32
31
30
HOsS~
H03S
90-51-7
CIoH9NO4S
HO
5639-34-9
CtoH9NO4S
CIoH9NO4S 116-63-2
SO3H
~
/OH
OH
ETAD (1990) Freeman (1987)
Chung (1981)
ETAD(1987) Kler (1986) Chung (1981) m + cyt
Gayathn (1980)
CRoHgNO 4384-92-3
120
119
119 C~oH,NO 2834-92-6 Wdlmms (1989)
DNA
m-
[contd]
Kter (1986) Dabney (1985) Garner (1977)
SAL SAL SAL
m mm-
f~
o
P~
e~
o
H
O
HO3S ~
.NH2
3
/
~
CIoH9NO7S2
S
SO3H OH
90-20-0
CIoHgNOTS2
OH
NI-12
SO3H
SO3H
OH
-
-
-
ETAD (1989)
Freeman (1987)
Chung (1980
Ames test data
m
DNA
m + GMA m- cyt
ETAD (1990)
Palmer (1979) Palmer (1979)
Other data
Table I--contd
122 C~0HgNO 606-41-7
CIoHgNO 2834-9 I-5
12l
119
No
test
Ames
Other data
Unsulphonated analogues
+ =posmve response - = negative response w = weak response ~ = quesUonnable results c =carcmogemoty, oral apphcatlon cell trans =cell transformation study haembmdmg= haemoglobin adducts m = genotoxmlty/mutagemcltym cyt = cytogemc~ty/chromosomemutation m DNA = DNA repair/indicator test m GMA = gene mutatton assay m eucaryotm cells (not Ames/Pnval) m SAL = Salmonella/mlcrosome (Ames) test pulm ad = pulmonary adenoma carcmogemclty test *With the exception of the Salmonella/mlcrosome assay, only a selecuon of the available data is shown For human carcinogens, only the IARC classlficaUon ts menUoned IARC classification. Group 1, carcmogemc to humans, Group 2B, possibly carcmogemc to humans, Group 3, not classifiable as to its carcmogemclty to humans
35
34
2007-20-7
CIoH9NO7S2
NH 2
33
H3OS
Structure
No
Aromatic ammosulphomc aods
z
Genotoxlclty and carcmogemcity of AASAs attempted to validate this test system using 65 chemicals together with available results of 2-year carcinogenicity and in vitro mutagemcity tests. The authors concluded that there was a poor agreement between genotoxicity results and the results obtained with strain A mice. Furthermore, results obtained with strain A mice are also not in agreement with results from 2-year carcinogenicity tests on aromatic amines and several other miscellaneous chemicals. Based on these findings, the concern of Theiss et al. expressed in their previous paper (Theiss et a l , 1981) about the possible carcmogenic potential of AASAs is certainly not justified. The aim of this paper ~s to investigate whether the available toxicity data on AASAs involved in dyestuffs chemistry give indications of any genotoxic or carcinogenic potential. The role of aromatic aminosulphonic acids as dyestuff intermediates and metabolites The sulphonated derivatives of the aminonaphthalenes and anilines are particularly important dye intermediates. The sulphonic acid group is used extensively in the dye chemistry to confer water solubility and for its ability to act as a good leaving group in nucleophllic substitutions. It is used almost exclusively for these purposes since it has only a minor effect on the colour of a dye. The amino group is the single most important functional group in dye chemistry. In azo-dye intermediates, it not only provides access to the azo group (by diazotization and coupling), but is also the most widely used auxochrome as can be seen from its presence in so many coupling components. Dye intermediates and similar compounds are an important group of primary metabolites of azo dyes, which constitute over 65% of all dyestuffs. Azo dyes all share one or several azo ( - - N : N - - ) groups, but they constitute a large variety of differing structures with a wide range of physlcochemical properties: from extremely water-soluble dyes to practically insoluble azo pigments, and from anionic and cationic to nonionic dyes. It has already been shown that under anaerobic conditions a variable percentage of the azo groups can be reduced enzymatically to the corresponding sulphonated and unsulphonated aromatic amines in bacteria and in mammalian systems including humans. In bis- and trisazo compounds the metabohc reduction of the azo groups proceeds stepwise under intermediate formation of the partly reduced molecules (i.e. azo-amino compounds). The formation of carcinogenic aromatic amines is assumed to be the major causative factor of the carcinogenic response to a given dye in an animal lifetime bioassay. Typical examples are the benzidine dyes, which are metabolized to the known human carcinogen benzidine. Based on present knowledge, it might be assumed that azo dyes, which are metabolized to a substantial
647
extent to carcinogenic amines in the living organlsms, should be suspected as potential carcinogens. Conversely, if non-carcinogenic amines are formed, the probabihty that the parent compound has carclnogenic properties becomes very small. It must be emphasized that beside this reductive cleavage of the azo group, other metabolic transformations could occur in the sense of additional activation or deactivation. Genotoxicity data of aromatic aminosulphonic acids As already mentioned, AASAs constitute a large number of potential metabolites especially of anionic dyes. Although few bioassay data on carcinogenicity exist, much information is available on genotoxiclty. This paper compiles a substantial amount of published and unpublished mutagenicity data. Table 1 displays available results of the Salmonella/microsomc test (Ames test), and other genotoxiclty and carcinogenicity data for 35 AASAs. For comparison, some available data on the unsulphonated analogues are also given. In contrast with mutagcnic and carcinogenic amines, such as 4-aminobiphenyl, benzidine and 2naphthylamine, the sulphonated analogues are not mutagenic in the Ames test. In the case of 2-naphthylamine several aminosulphonic acid isomers were non-mutagenic (Le et al., 1985). In a recent publication (Freeman et al., 1987), the results from the Ames and Prival mutagenlcity test of six aminonaphthalenesulphonic acids were reported. Five of them showed a very weak positive response (i.e. 1amino-naphthalene-7-sulphonic acid, 1-amino-8-hydroxynaphthalene-4-sulphomc acid, 2-amino-6-sulphonic acid (24), 2-amino-8-hydroxynaphthalcne-6sulphonic acid 32 (g-acid) and 3-ammo-8-hydroxynaphthalene-6-sulphonlc acid (y-acid)), whereas l-amlno-8-hydroxynaphthalene-3,6-disulphonic acid (34) was negative in both tests. The authors presume that the very weak positive response exhibited by the five amlno-naphthalenesulphonlc acids might be due to impurities since the samples used in their assays were not 100% pure. Such an assumption is supported by tests with highly purified materials. Le et al. (1985) found 2-aminonaphthalene-6-sulphonic acid (24) to be non-mutagenlc in the Salmonella/ microsome test in the presence of hepatic S-9 derived from the rat, hamster, monkey and humans. In the same test, using S-9 derived from the rat, 2amlno-8-hydroxynaphthalcne-6-sulphonic acid (32) (),-acid) (ETAD, 1990) and very recently 3-amino8-hydroxynaphthalene-6-sulphonic acid (J-acid) (ETAD, 1991) gave a negative response (not listed in Table 1). Similar experiences regarding purity were encountered with 2-amino-5-methylbenzenesulphonic acid (5) (4B-acid) and 2-amino-5-chloro-4-methylbenzenesulphonic acid ( l l ) (CLT-acid), which were negative in the Ames test only after thorough
No
HO3S -
-
~
C6HTNSO3
-
Structure
NH 2
Aromattc aminosulphomcacid (hypothetical dye metabohte)
N~N
C.I. 19140 C.I. Acid Yellow 23
NaO3S---~
C.l. Food Yellow 3
C.I. 15985
NaO3S~
C.I, 14600 C.I. Acid Orange 20
SO3Na
COONa
HO ~-----N~SO3Na
HO
Borzelleca (1988a) Borzelleca (1988b) Maekawa (1987)
IARC (1987) NTP-TR-208 (1981)
BIBRA (1988)
Azo dye wRh negaUveresults m caremogemcRyassays IARC (1987) Schach von WIttenau (1983)
Table 2. Ammes from azo dyes with negatwe results m caranogematy assays*
I01
No.
Unsulphonated analogue of amine
l0
CH30
/~.~-"-
. CH3
CaHItNSO 4
HO3S
T
NH~
CoHTNSO3
H03S
NH2
HO
N= N
C.I. 16035 C.I. Food Red 17
NaO3SH 3 C ~
cH3
OCH 3
C.I. Food Red 1
C.L 14700
NaO3S
C.I. 13065 C.I. Acid Yellow 36
NaO3S~__
C.I. 15980 C.I. Food Orange 2
Na03S
HO
NaO3S
HO
SO3Na
NH©
SO3Na
Borzelleca(1989)
IARC (1987) Schach von Wittenau (1983)
Vettoraz~ (1981) ADL (1980)
Vettorazzl (1981)
[contdl
107
101
> >
o
=s
o
21
No
C10H9NSO3
SO3H
NH 2
Structure
AromaUc amlnosulphomcacid (hypotheUcal dye metabohte)
C.I. 20285 C.I. Food Brown 3
NaO3S
N~N
SO3Na
OH
N~N
HO
N~N
C.I. 14720 C.I. Acid Red 14
C.I. 16185 C.I. Acid Red27
NaO3S~
SO3Na
N~-----NS
~
SO3Na
SO3Na
C~OH
HO
HO
SO3Na
Vettorazzi (1981) Drake (1978) Carpamm (1978)
BIBRA (1990) NTP-TR-220 (1982)
ADL (1980) Clode (1987)
Azo dye with negative results m carcmogenlcRyassays
Table 2 - - - c o n t d
107
No
amine
Unsulphonated analogue of
o~
37
36
CsHIINSO3
c~
~ C ~ s 0 3
NH2
CsHIINSO3
CH3
C H 3 ~
NI-I2
H
SO3H
NaOsS
i~c
NaO3S
CH3
N
O3Na ~
C.I. 14700 C.I. Food Red 1
c~
HO
~
NaO3S
N
HO
C.I. 16255 C.I. Acid Red 18
C.I. 14815 C.I. Food Red 2
NaO3S
SO3Na
HO
SO3Na
IARC(1987) SchachyonWRtenau(1983)
Vettorazzl(1981)
Brantom(1988) BIBRA(1986) Mason(1974)
[conta]
123
123
~
r.~
0
g~ e~
o
g
38
!1
NO.
HO3S
HOsS
HOsS
"CH3
C IoHIoN2SO3
NH 2
CTI'IgNSOs
CHs
CTI~NSO3CI
Cl
N~
Structure
Aromattc amlnosulphonicacid (hypothetical dye metabohte)
N
HO
NaO3S---O
C.I. Food Black 1
SO3Na
COONa
NaO3S
HO
SO3Na
NHCOCH 3
BIBRA (1989b) Vettorazzl (1981) Drake (1977) Gaunt (1972)
BIBRA(1982a)
BIBRA (1989a) IARC (1987) NTP-TR-225 (1982)
Azo dye with negattve results in carcinogemcttyassays
C.I. 28440
C.I. 15850 C.I. Pigment Red 57
N
O3Na
C.I. Pigment Red 53:1
C.I. 15585:1
N
SO3Na
CHs--~
CI
OH
Table 2---contd
124
102
No 108
2nllne
Unsulphonated analogue of
Z
CIoH9NSO4
H O 3 S / ~
NH2
OH
t
C.I. 16035 C.I. FoodRed 17
H3C
C.I. 15985 C.I. FoodYellow3
NaO3S ~N'---N ~
HO
N_~_NHO~
C.I. 15980 C.I. FoodOrange2
NaO3S~
SO3Na
SO3Na
SO3Na
Borzelleca (1989)
BIBRA (1988) IARC(1987) NTP-TR-208(1981)
Vettorazzt(1981)
[cored]
!19
r~
>.
o
o
33
No. 39
HO3S
CI0HgNS207
NH2
CIoHgNS207
HO3S ~ O H
Structure H03S
Aromaticammosulphomcacid (hypotheucaldye metabohte)
SO3H
NaO3S
NaO3S - - ~
N = . N - ~
C.I. 16255 C.I. Acid Red 18
N~ N
C.I. 16185 CT Acid Red 27
N
N
SO3Na
S03Na
SO3Na
HO SO3Na
C.I. 16230 C.I. Food Orange 4
Na03S
HO
IARC (1987) Clode (I987)
Zeiger (1987) NTP-TR-211 (1987) IARC (1987)
Brantom (1987) BIBRA (1986) Mason (1974)
Azo dye with negativeresults m carcinogenicttyassays
Table 2 - - c o n t d
119
119
No.
Unsulphonated analogue of amine
o~
41
40
j,.~ NH2
/NH 2
CIoH9NSO4
SO3H
~
OH
C toH9NS04
SO3H
OH
NaO3S
N
C.I. 14700 C.I. Food Red 1
C.I. 14815 C.I. Food Red 2
• SO3Na
HO
N
HO
NaO3S
C.I. 14720 C.I. Acid Red 14
SO3Na
OH ~
SO3Na
SO3Na
Vettorazzl (1981)
Schachyon Wittenau(1983)
IARC (1987)
BIBRA(1990) NTP-TR-220(1982)
[contd]
119
119
u.
o'J
o
o
o
HO3S
SO2I-IN. OH
OH
CloH1oN2S207
H2N
C12HI4N2S20s
SOsH
CITHI6N209S 3
H O s S / ~
H,C----~
Structure
So3NH2H
©
©
~
~
OH
NH2
C.I. 17200 C.I. Acid Red 33
NaOsS
N~N
N
SOsNa
SO3Na
NHCOCH3
C.I. 18050 C.I. Food Red 10
NaO3S
N
HO
~
SO3Na
_ NHO2S
C.I. Acid Red 106
C.I. 18110
NaOsS
N~-N ~
HO
~ CI-I~
BIBRA(1982b) BIBRA(1982c)
VettorazzJ(1981) WHO(1977)
IARC/WHO(1981)
Azo dye withnegativeresultsin carcmogemcltyassays
127
126
125
No
amine
Unsulphonated analogueof
*ForaromaUcammosulphomcacids withnumbers 1-35and for unsulphonatedanalogueswithnumbers101-122,see Table 1 (mutagemcltyand othertoxicitydata of selectedaromaUcammosulfomcacidsand their unsulphonatcdanalogues).
44
43
42
No.
Aromattcammosuiphomcacid (hypotbeucaldye metabohte)
Table 2--contd
Z e~
657
Genotox~clty and carcmogeniclty of AASAs purification. Both substances were also negative in the micronucleus assay (ETAD, 1988). The two compounds reported by Freeman et al. (1987) to be weakly positive m the Ames test (i.e. lamino-naphthalene-7-sulphonic acid and 1-amino-8hydroxynaphthalene-4-sulphonic acid) are not listed as mutagemc in Table 1 of this review in view of the uncertainty regarding the purity of the samples tested. Unlike benzidme (115) and 2-aminonaphthalene (118), aniline (101) and p-ammotoluene (102) have only weak (aniline) or no significant carcinogenic potential m animal studies and are not mutagemc in the Ames test. The sulphonated analogues of aniline and p-aminotoluene are therefore, as expected, not mutagenic in the Ames test. In some cases, methyl and chlorine subst~tuents m the amline molecule considerably influence the genotoxic or carcmogemc potency. Several of these chemicals are mutagenic m the Ames test. The sulphonated derivatwes, however, as shown in Table 1, were not mutagenic m the Ames test. In addition to the results from the Ames test, some of the compounds gwen in Table 1 have been studied in other tests on genotoxiclty (m vivo and m vitro) including cell transformation assays. The outcome of all these studies reflects, with only few exceptions, the absence of genotoxicity for AASAs. 2-Ammonaphthalene-1,5-disulphonic acid (26) and the disulphonlc acid derivatives of benzldine, (18) and (19), were not active in cell transformation assays m vitro, whereas the unsulphonated carclnogemc amines induced cell transformation m vitro In contrast to 2-aminonaphthalene (118), which has been reported to cause D N A damage (positwe m several UDS tests), the l-sulphonic acid analogue (22) was negatwe in th~s test system. 2-Aminonaphthalene induced gene conversion in Saccharomyces cerewsiae with and without metabolic activation. 2-Naphthylamine-l-sulphonic acid (22) did not produce gene conversion m trp or ade genes of S. cerevistae strata D4 (Della Porta et al., 1982). Supportive evidence of non-genotoxlclty for AASAs also comes from mutagenicity studies with azo dyes usmg reductive assay conditions. Structures of Direct Black 19 analogues with unsulphonated p-phenyleneblsazo moieties were active m the Salmonella/microsome assay, whereas the sulphonated derivatives were not (Lm and Solodar, 1988). To summarize the results, in general aromatic phenyl- and naphthyl-aminosulphonic acids are nonmutagenic to Salmonella. Other tests on genotoxiclty have also been considered. The data compiled show no or very low mutagenic potentml. In addition, three AASAs were negative in feeding studies on carcmogenicity. The available data indicate that sulphonation of phenyl- and naphthylamines decreases toxicity considerably. Where comparable data on sulphonated and unsulphonated analogues exist, loss
of genotoxlc and/or carcinogenic suiphonation is observed.
activity
on
Mechanistic considerations It has been repeatedly reported (Combes and Haveland-Smith, 1982; Garner and Nutman, 1977) that sulphonat~on renders amines less mutagenic and non-carcinogemc, and azo dyes based on AASAs non-carcinogenic (see Table 2). It appears to be a general rule that highly sulphonated azo dyes are poorly absorbed from the gut (Walker, 1970), and therefore become b~oavailable to a significant extent only after azo reducuon to the corresponding ammosulphonic acids. The degree of absorption from the gut is dependent on the polarity of the molecule, and sulphonation increases the polarity of the molecule For sulphonated amines, significant absorption has been observed (for review see Walker, 1970). Monosulphonated amines (e.g. sulphanilic acid, naphthlomc acid) are absorbed to a large extent, possibly m the zwitterionic form. Unlike aromatic amines, however, ring substitution by sulphonation of azo dyes based on AASAs prevents the activation of amino groups by liver metabolism (Garner and Nutman, 1977). Lin and Solodar (1988) beheve that sulphonation (as in the case of p-phenylenediamine) Increases the electronegatlvity of the metabolized molecule and thus decreases the electrophilicity required to attack D N A bases. Recently, Levme (1991) m h~s review on metabohsm of azo dyes explained that highly charged sulphonated azo dyes resist bacterial enzymic attack, and are poorly absorbed from the intestinal tract thus providing poor access to the hver endogenous metabolism As m the case of the non-carcmogemc, non-mutagenic 2-naphthylammo-l-sulphomc acid, pharmacokmetic studies indicate a rapid elimination of unchanged aminosulphonic acid after oral or iv administration. No biotransformation or cleavage of the sulphonic group was detected (Della Porta, 1982). As a more general conclusion to a study on the structural bas~s of the mutagemc~ty of phenylazoanihne dyes, Rosenkranz and Klopman (1989) reported that sulphonation appears to interfere with a structure in the azo dyes that is required for oxidation of the amino group A negatwe inductive effect of the sulphomc group may influence azo reduction or amine oxidation Combes and Haveland-Smlth (1982) suggested that the sulphonlc group may exert an unfavourable electromc influence on the activation of the 4-amino-Natom by exerting a strong negative inductive effect. The negative inductive effect of the sulphonic acid group on azo reduction has been demonstrated; Walker and Ryan (1971) showed that analogues of Red 2G and Red 10B sulphonated in the p-position of the phenyl ring were reduced by cell-free extracts
658
R. JUNG et al.
of Streptococcus faecahs two to three times more rapidly than the parent compound, owing to the substitution with an electron-withdrawing group.
Absence of evidence of the carcinogenic potential of aromatic aminosulphonic acids Few carcinogenicity studies have been carried out on substituted amlnonaphthalene- and aminobenzenesulphonic acids, probably because these compounds are expected to be non-carcinogenic. However, the existence of a considerable number of negative bioassays of azo dyes, which are metabolized to such aods, give additional evidence of their low chronic toxicity and their non-carcinogenic (or at most low carcinogenic) potential. It has been demonstrated that azo dyes are metabolized in the organism to aromatic amines by reductive cleavage of the azo group in bacterial systems both in animals and humans. However, the rate of this reaction is very much dependent on the individual structure and can differ widely. This has been shown by Watabe et al. (1980) in their study on the reduction of four azo dyes by micro-organisms from human faeces. The formation of AASA metabolites is certainly not always quantitative. There are several factors influencing the rate of formation such as other competing metabolic processes, rate of absorption, route of admimstratlon, low capacity of the reductlve enzymes, lack of availability of the dye to these enzymes, steric hindrance, etc. Table 2 lists azo dyes for which adequate negative 2-year bioassays have been done. With a few exceptions, the dyes can hypothetically form only AASAs (also compiled in Table 2) on reductive cleavage of the azo group. Considering the very high doses applied in most of the studies shown in Table 2, it can be assumed that substantial quantities of the corresponding AASAs have been formed in the gastro-intestinal tract and/or in the liver. The lack of tumorigenic response in these studies is indicative of a very low or even absent carcinogenic potential of these metabolltes. This conclusion is supported by the genotoxicity data summarized in Table 1. It is generally accepted that genotoxicity is a primary event in the carcinogenic process. Therefore the absence of genotoxiclty with amlnosulphonic acid supports the conclusion that these compounds do not act as genotoxlc carcinogens. Table 1 also includes negative carcmogeniclty bioassay data for several aminosulphonic acids. In the case of benzidine and 2-naphthylamine, it has been shown experimentally that sulphonation of the parent carcinogenic amine leads to non-carcinogenic substances. Although there is a long history of production of a large number of AASAs in the dyestuff manufacturing Industry, there are no indications of any carcinogenic risks from these compounds. In
contrast with these findings, bladder cancer cases resulting from exposure to 2-naphthylamine and benzidine are well known. An interesting observation in this connection was made by Kumar et aL (1981), who found in their immunological studies a correlation between increased lymphocyte reactivity and bladder cancer risk in workers exposed to 2-naphthylamme (118). Workers exposed to 2-naphthylamine-l-sulphonic acid (22), but not to 2-naphthylamine, had lymphocytes with a normal range of reactivity, which is in keeping with the suggestion that this compound is non-carcinogenic.
Conclusion The evaluation of available information and data on AASAs, either in use for dyestuff manufacture or being formed by reductlve cleavage of azo dyes, shows that these compounds, unlike some known carcinogenic aromatic amines, generally have no or very low genotoxic effect and hence no or minimal tumorigenic potential. In addition, considering the low exposure to AASAs prevailing in the techmcal application and use of azo dyes, there is not likely to be any significant toxic risk if AASAs are formed by metabolic cleavage in the organism or in the environment, or occur as micro-contaminants in colourmgs.
REFERENCES ADL (1980) Arthur D LRtle Report Azo dyes: evaluation of data relevant to human health and environmental safety. Anhker R (1977) Color chemistry and environment Ecotoxwology and Environmental Safety 1, 60-72. Anliker R. (1983) Die orgamschen Farbstoffe und Pigmente lm Spannungsfeld der Umwelt und Gesetzgebung. Textilveredlung 18, 92-98 Ashby J., Paton D., Lefevre P. A., Styles J. A. and Rose F. L. (1982) Evaluation of two suggested methods of deactivating orgamc carcinogens by molecular modification Carcmogenests 3, 1277-1282. Ashby J, Styles J. A and Paton D (1978) In vRro evaluation of some denvaUves of the carcinogen Butter Yellow lmphcatlons for environmental screening. British Journal of Cancer 38, 34-50. BIBRA (1982a) Four D & C Reds permanently listed. BIBRA Bulletin 21, 558 BIBRA (1982b) No adverse effects from D & C Red No. 33. BIBRA Bulletin 21, 63. BIBRA (1982c) Reports on three D & C colourmgs BIBRA Bulletin 21, 233 BIBRA (1984) Chemical Hazard Information Profile on ammophenols. B1BRA Bullet,n 23, 196 BIBRA (1986) ToxicRy profile of Ponceau 4R BIBRA (1988) ToxicRy profile of Sunset Yellow FCF. BIBRA (1989a) Toxicity profile of D & C Red 9. BIBRA (1989b) Toxioty profile of Black PN. BIBRA (1990) Toxicity profile of carmmsme Birner C and Neumann H-G. (1988) Biomomtonng of aromatic amines II. hemoglobin binding of some monocyclic aromatic amines Archwes of Toxicology 62, 110-115
Genotoxiclty and carcinogemcity of AASAs Borzelleca J. F. and Hallagan J. B. (1988a) Chronic toxicity/ carcmogemcity studies of FD & C Yellow No. 5 (tartrazine) in rats Food and Chemical Toxicology 26, 179-187. Borzelleca J. F and Hallagan J. B (1988b) A chronic toxiclty/carcmogemclty study of FD & C Yellow No 5 (tartrazme) in mice. Food and Chemical Toxwology 26,
189-194 Borzelleca J F , Olson J. W and Reno F E (1989) Lifetime toxJclty/carcmogemcity study of FD & C Red No. 40 (Allura Red) m Sprague-Dawley rats Food and Chemical Toxicology 27, 701-705. Brambdla G , Carlo P , Fmollo R and Ledda A. (1985) Vlscometnc detection of hver DNA fragmentation in rats treated with ten aromatic amines Discrepancies with results prowded by the alkahne elution technique. Carcinogenesis 6, 1285-1288. Brantom P. G , Stevenson B I. and Wright M G (1987) Long-term toxicity study of Ponceau 4R in rats using ammals exposed m utero Food and Chemical Toxicology 25, 955-962. Carpanml F M B, Butterworth K. R , Gaunt I F., Kiss I S, Grasso P and Gangolh S D (1978) Longterm toxicity studies on Chocolate Brown HT m rats Toxwology 11, 303-307 Cesarone C F., Bolognesl C. and Santl L. (1982) Evaluation of damage to DNA after in vwo exposure to different classes of chemicals Archwes of Toxicology (suppl 5) 355-359 Chung K. T . Fulk G E and Andrews A W (1978) The mutagenlcity of Methyl Orange and metabohtes produced by intestinal anaerobes. Mutation Research 58, 375-379 Chung K T., Fulk E F and Andrews A W (1981) Mutagemcity testing of some commonly used dyes Apphed and Environmental Microbiology 42, 641~48 Clode S A , Hooson J., Grant D. and Butler W. H (1987) Long-term toxicity study of amaranth in rats using ammals exposed m utero Food and Chemwal Toxicology 25, 937-946. Combes R. D. and Haveland-Smlth R. B (1982) A review of the genotoxiclty of food, drug and cosmetic colours and other azo, trlphenylmethane and xanthene dyes. Mutatwn Research 98, 101-248 Dabney B J , DeBroy J. A., Freeman H. S, Price K. B, Esancy J., Calogero F , Kennedy K M and Whaley W. M (1985) Structure-actwlty relationship for azo dyes made from aminonaphthalenes. Poster, USOC No 489 De Flora S., Zanacchl P., Camolrano A., Benicelh C and Badolatl G S (1984) Genotoxlc activity and potency of 135 compounds in the Ames reversion test and m a bacterial DNA-repair test Mutation Research 133, 161-198. Della Porta G and Dragam T A. (1982) Noncarclnogemcity in mice of a sulfomc acid derivative of 2-naphthylamme. Carcmogenests 3, 647~49. DETO (1980) Dyes Environmental and Toxicology Orgamzation, lnc Toxicology Summary Dlenckx P J (1989) Cytotoxicity testing of 114 compounds by the determination of the protein content in HEP G2 cell cultures. Toxicology m Vitro 3, 189-193 Drake J J -P, Butterworth K. R , Gaunt I F. and Grasso P (1977) Long-term toxicity study of Black PN in mice Food and Cosmetics Toxicology 15, 503-508. Drake J. J -P, Butterworth K. R , Gaunt I. F. and Hardy J (1978) Long-term toxicity studies of Chocolate Brown HT in mice. Toxzcology 10, 17-27 Ennever F. K. and Rosenkranz H S. (1986) Predicting the carcmogemc~ty of the aromatic amine denvatwes tested in the second UKEMS collaborative study. Mutageneszs 1, 119-123
659
ETAD (1981-1991) Ecological and Toxicological Association of Dyes and Organic Pigments Manufacturers Various individual ETAD internal reports of unpubhshed studies by ETAD Member Companies Fassma G., Abbondandolo A , Mariam L., Tanlngher M. and Parodi S. (1990) Mutagemcity m V79 cells does not correlate with carcmogemty in small rodents for 12 aromaUc amines. Journal of Toxicology and Environmental Health 29, 109-130. Freeman H. S., Esancy J F., Esancy M. K , Mdls K. P. and Whaley W M (1987) An approach to the design of nonmutagemcazodyes I Theldentlficationofnonmutagemc precursors and potential metabolltes Dyes and Pigments 8, 417-430. Garner R C , Martin C N. and Clayson D. B (1984) Carcmogenlc aromatic amines and related compounds. In Chemical Carcmogens Vol. 1 2nd Ed. Edited by Ch E. Searle ACS Monograph 182 Garner R C and Nutman C A (1977) Testing of some azo dyes and their reduction products for mutagemclty using Salmonella typhimurtum TA1538. Mutation Research 44, 9-19 Gaunt I, F., Carpanml F. M , Grasso P., Kiss I S. and Gangolh S D (1972) Long-term feeding study on Black PN m rats. Food and Cosmetics Toxicology 10, 17-27. Gayathn M V and K~shnamurthy N B (1980) Prehminary studies on toxicity and mutagenlclty of 1-ammo-2naphthol-4-sulfomc acid m Drosophila melanogaster. Journal of Biosclence 2, 49-54 Harrison J H. and Jollow D J (1987) Contribution of aniline metabohtes to amhne-mduced methemoglobmemia Molecular Pharmacology 32, 423-431. IARC (1987) Suppl. 7 Overall Evaluations of Carcmogemclty: an updatmg of IARC Monographs Volumes 1-42 International Agency for Research on Cancer, Lyon IARC/WHO (1981) Information Bulletin on the survey of chemicals being tested for carcmogenicity No. 9, 60-61 Ishldate M , Harnois M. C. and Sofuni T. (1988) A comparative analysis of data on the clastogemcity of 951 chemical substances tested in mammahan cell cultures Mutation Research 195, 151-213 Kler L E , Bruslck D. J , Auletta A. E , Von Halle E. S, Brown M, M , Simon V. F , Dunkel V , McCann J , Mortelmans K., Prlval M., Rao T. K and Ray V (1986) The Salmonella typhtmur,um/mammahan microsomal assay--a report of the U.S Environmental Protection Agency Gene-Tox Program Mutatzon Research 168, 69-240 Kirchner G and Bayer U. (1982) Genotoxlc actiwty of the amlnophenols as evidenced by the induction of sister chromatld exchanges. Human Toxicology 1, 387-392 Kumar S, Taylor G., Hurst W., Wilson P and Costello C B. (1981) Lymphocyte reactivity of workers exposed to carcmogemc and noncarcmogemc chemicals Brutish Journal of Industrial Medicme 38, 167-169 Le J., Jung R. and Kramer M. (1985) Effects of using liver fractions from different mammals, including man, on results of mutagemcity assays m Salmonella typhtmurmm. Food and Chemtcal Toxicology 23, 695-700 Levine W G. (1991) Metabolism of azo dyes: implication for detoxicatlon and actwat~on Drug Metabohsm Reviews 23, 253-309 Lm G. H. Y and Solodar W E. (1988) Structure activity relationship studies on the mutagemcity of some azo dyes in the Salmonella/microsome assay. Mutagenests 3, 311-315. Maekawa A , Matsouka C , Onodera H , Tanigawa H., Furuta K., Kanno J., Jang J J , Hayashl Y and Ogre T (1987) Lack of carcmogemclty of tartrazme (FD & C Yellow No 5) in the F344 rat. Food and Chemical Tox,eology 25, 891-896.
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Maronpot R. R., Shimkln M. B , Wltschi H P, Smith L H. and Chne J M. (1986) Strain A mouse pulmonary tumor test results for ch~nicals previously tested in the National Cancer Institute Carcinogenlcity tests. Journal o f the National Cancer Institute 76, 1101-1112. Mason P. L , Gaunt I. F , Butterworth K R and Hardy J. (1974) Long-term toxicity of Ponceau 4R in mice BIBRA Bulletin 13, 343-344. Mitchell A. D (1988) Evaluation of the L5178Y mouse lymphoma cell mutagenesls assay, methods used and chemicals evaluated. Enwronmental and Molecular Mutagenests 12 (suppl. 13), 1-18 Neumann H.-G (1988) Bmmomtoring of aromatic amines and alkylating agents by measuring hemoglobin adducts International Archives o f Occupatwnal and Enwronmental Health 60, 151-155 NTP-TR-36 (1978) Carcinogenesis bloassay of o-anthramhc aod National Toxicology Program Technical Report 36 NTP-TR-142 (1978) Carcinogenesis bloassay of p-cresidine. National Toxicology Program Technical Report 142. NTP-TR-145 0978) Carcinogenesis bioassay of 3-chlorop-toluldIne National Toxicology Program Technical Report 145. NTP-TR-208 (1981) Carcinogenesis bIoassay of FD & C Yellow No. 6. National Toxicology Program Technical Report 208. NTP-TR-220 (1982) Carcinogenesis bmassay of Acid Red 14 National Toxicology Program Technical Report 220 NTP-TR-225 (1982) Carcinogenesis bloassay o f D & C Red No 9. National Toxicology Program Techmcal Report 225 NTP Rep. (1984) Chemical test results for mutagemcity In Salmonella assays in FY 1983 National Toxicology Program Reports NTP Rep 85-002 (1985) Fourth Annual Report on Carcinogens. National Toxicology Program Reports NTP Rep. (1985a) Chemical test results for mutagemclty in Salmonella assays m FY 1984 Natmnal Toxicology Program Reports NTP Rep. (1985b) Chemical test results for cytogenetic effects in Chinese hamster ovary cells m FY 1984. National Toxicology Program Reports. NTP Rep (1986) Chemical test results for mutageniclty in Salmonella assays in FY 1985 National Toxicology Program Reports. NTP-TR-211 (1987) Carcinogenesis bloassay of Acid Orange 10. National Toxicology Program Techmcal Report 211. NTP Pep (1988) Chemical test results for cytogenetlc effects m Chinese hamster ovary cells m FY 1987 National Toxicology Program Reports. NTP Rep. (1989a) Chemical test results for mutagemclty In Salmonella assays in FY 1988. NTP Rep. (1989b) Chemical test results for cytogenetic effects in Chinese hamster ovary cells In FY 1988. National Toxicology Program Reports Pai V., Bloomfield S. F. and Gorrod J. W (1985) Mutagenicity of N-hydroxylamlnes and N-hydroxycarbamates towards stratus of Eschertchta coh and Salmonella typhtmurtum. Mutatmn Research 151, 201-207 Palmer K. A , Sheu C. W. and Green S. (1979) Mutagemcity study of R-amino salt A metabolite of Amaranth (FD & C Red. No. 2), in mouse lymphoma cells heterozygous at the thymidine klnase locus and in the rat dominant lethal test. Food and Cosmencs Toxzcology 17, 5-9. Pogodina O N., Svetlova M. P., TomIhn N. V , Phss G. B. and Khudolel V. V. (1984) Correlation of mutagenic and carcinogenic activities of certain aromatic amines. Eksp Onkol. 6, 23-25.
Purchase I. F H , Longstaff E., Ashby J., Styles J. A., Anderson D., Lefevre P. A. and Westwood F. R. (1978) An evaluation of 6 short-term tests for detecting organic chemical carcinogens. Britssh Journal of Cancer 37, 873-959. Rosenkranz H. S and Klopman G. (1989) Structural basis of the mutagemclty of phenylazoanlline dyes. Mutanon Research 221, 217-234. Schach von Wittenau M. and Estes P. C (1983) The redundancy of mouse carcInogenlclty bioassays. Fundamental and Apphed Toxicology 3, 631-639. Shahin M. M (1989) Studies on the mutageniclty of aniline in association with Norharman In the Salmonella/ mammalian mlcrosome assay. International Journal o f Cosmetzc Science 11, 129-140 Shimlzu H., Suzuki Y., Takemura N , Goto S. and Matsusbita H. (1985) The results of microbial mutation test for forty-three industrial chemicals Japanese Journal o f Industrial Health 27, 400-419 ShlmlZU H and Takemura N (1983) Mutagemcity of some aniline derivatives 1lth meeting on Occupational Health in the Chemical Industry Proceedings of the International Congress In 1983 497-508 Styles J. A (1973) Mammalian cell transformation m wtro. Britzsh Journal o f Cancer 37, 931-936 Thelss J. C , Shlmkln M B and Weisburger E K. (1981) Pulmonary adenoma response of strain A mice to sulfonlc acid derivatives of 1- and 2-naphthylamines Journal of the National Cancer Institute 67, 1299-1302. Valmo H , HemminkI K and Wilbourn J. (1985) Data on the carclnogemclty of chemicals in the IARC monographs programme Carcinogenesis 6, 1653-1665 Vemtt S and Bushell C. T (1976) Mutagemcity of the food colour Brown FK and constituents in Salmonella typhimurmm Mutation Research 40, 309-316. Vettorazzi G. (1981) Handbook of Internatmnal Food Regulatory Toxzcology Vol. 2' Profiles. MTP Press, Lancaster, UK Von der Hude W , Behm C , Guertler R and Basler A (1988) Evaluation of the SOS chromotest. Mutation Research 203, 81-94 Walker R (1970) The metabohsm of azo compounds, a review of the literature Food and Cosmetics Toxicology 8, 654-676. Walker R. and Ryan A J (1971) Some molecular parameters influencing rate of reduction of azo compounds by intestinal mIcroflora Xenobtotica 1, 483-486 Wangenhelm J. and Bolcsfoldl G (1988) Mouse lymphoma L5178Y thymldme klnase locus assay of 50 compounds. Mutagenests 3, 193-205 Watabe T , Ozawa N , Kobayashl F and Kurata H. (1980) Reduction of sulphonated water-soluble azo dyes by micro-organisms from human faeces Food and Cosmetics Toxicology 18, 349-352 WHO (1977) Red 2G' evaluatmn of acceptable daily Intake World Health Organization Food Additives series no 12, 94-102 Wilhams G M. (1989) Structure-activity relationship in the rat hepatocyte DNA-repair test for 300 chemicals. Mutanon Research 221,263-286 Yoshiml N , Sugle S, Iwata H , Ntkwa K.. Mort H, Hashlda C. and Shimizu H (1988) The genotoxlcIty of a variety of amhne derivatives in a DNA repair test with primary cultured rat hepatocytes. Mutation Research 206, 183-191 Zeiger E (1987) Carcinogenlcity of mutagens. Predictive capability of the Salmonella mutagenesis assay for rodent carcinogemclty Cancer Research 47, 1287-1296 Zeiger E , Anderson B., Haworth S, Lawlor T and Mortelmans K. (1988) Salmonella mutagenmty tests IV Results from the testing of 300 chemicals. Enwronmental and Molecular Mutagenesls 11, 1-158
Fd Chem. Toxw Vol 30, No. 7, pp 635-660, 1992 Pnnted m Great Britain All rights reserved
Review Section A COMPILATION OF GENOTOXICITY AND CARCINOGENICITY DATA ON AROMATIC AMINOSULPHONIC ACIDS R. JUNG, D. STEINLE* and R. ANLIKER* Hoechst AG, Gewerbetoxlkologle, PO Box 800320, D-6230 Frankfurt am Main, Germany and *Ecological and Toxicological Assocmtlon of Dyes and Orgamc Pigments Manufacturers (ETAD), PO Box, CH-4005, Basel, Switzerland (Accepted 16 March 1992)
Summary--A review is presented to evaluate existing information on genotox~city and carcinogenicity testing of various aromatic aminosulphomc acids (AASAs) A great variety of water-soluble azo dyes can form aromatic phenyl- or naphthyl-ammosulphomc acids by ebemlcal and enzymatic reduction. AASAs are also used as intermediates in the synthes~sof azo dyes and azo p~gments and can anse as contaminants in the final products Comparisons have been made with the data available on the corresponding unsulphonated analogues, some of which are known to be genotox~c and/or carcinogemc The vast majority of the AASAs were concluswely non-mutagenic in the Ames test. In most cases the absence of genotoxic~tywas also demonstrated with a variety of other test systems m wtro and m vivo It is concluded that AASAs, m contrast with some of their unsulphonated analogues, generally have no or very low genotoxtc and tumorigemc potentml.
Introduction Epidemiological data have shown an association between occupational exposure to 2-naphthylamme (no. 118 in Tables), benzidme (115), 4-aminobiphenyl (114) and 2-methyl-4-chloroaniline, and the risk of cancer of the urinary bladder. The carcmogenicity effects in humans and animals of a number of lipophilic aromatic amines are well documented, and experimental evidence indicates that this carcinogenicity is dependent on metabolic actwation. On the other hand, several aromatic amines have been shown not to possess tumorigenic actwity even when they are positive in the Ames test, clearly indicating that carcinogenicity is not a common feature for all compounds in this class of chemicals. Aromatic amines are important intermediates m the synthesis of a great variety of products, such as plastic materials, pesticides, rubber chemicals, and colourings, primarily azo colourings. The production and use of the above-mentioned four carcinogenic amines has been abandoned by the major dyestuffs manufacturers, and in many industrialized countries their production and use are controlled by very stringent regulations. 2-Naphthylamine (118) also played an important role as a starting material in the synthesis of key intermediates for colourings, especially aminonaphthalenesulphonic acids. *To whom correspondence should be addressed. Abbreviation: AASA = aromatic armnosulphomc acid.
Alternative synthetic routes to these aminonaphthalenesulphonic acids were developed to circumvent the use of the hazardous 2-naphthylamine (Anliker, 1977). Undoubtedly, substituted aminonaphthalenesulphonic acids, and generally aromatic aminosulphonic acids (AASAs; often also called 'sulphonated aromatic amines') will continue to be major intermedmtes in the manufacture of organic colourings. The seeming structural similarity between AASAs and the unsulphonated analogues gave nse to the suspicion that they might also possess a similar tumorigemc activity. As in other classes of chemicals, some of the existing shortcomings of structure/activity analogies can also be shown with these groups of chemicals (Anliker, 1983). The sulphonic acid closely related to 2-naphthylamine (118), 2aminonaphthalene-1-sulphonic acid (22), did not produce any carcinogenic effects in a chronic feeding study in mice, even at high dose levels of 1000 mg/kg body weight (Della Porta, 1982). Theiss et al. (1981) stud~ed seven 1-amino- and 2-aminonaphthalenesulphonic acids in the lung adenoma test in the strain A mouse using lp administration. The authors concluded that neither l-naphthylamine (117) nor the four sulphonic acid derivatives of l-naphthylamine tested were tumorigenic. However, 2-naphthylamine and two out of the three sulphomc acid derivatives of 2-naphthylamine tested were believed to have produced a statistically significant tumour response. A later paper by the same group (Maronpot et al., 1986) 635
N~
I
88-21-1
C6HTNO3S
~
121--47-I
C6HvNO3S
SO3H
~ S 0 3 H
N~
121-57-3
C6H~NO3S
so3H
+
Structure
No.
Zetger (1988) NTP Rep (1986) Shimtzu (1983)
Zelger (1988) NTP Rep (1986) Chung (1981) Chung (1978) Vemtt (1976)
ETAD (1989) + (w) Zeiser (1988) + (w) NTP Rep (1986)
-
-
Ames lest data
Aromatic ammosulphonic acids
m
DNA
m - cyt
Yoshtml (1988)
Dienckx (1989)
Other data 101 C6HvN 62-53-3
No. -
test
Ames
Group 3
C+
m+ c+ m+ mmm+ m+ m c+
Other data
GMA haembmding cyt cyt DNA GMA GMA DNA haembmdiug
m - SAL m - SAL m - SAL
Unsulphonated analogues
Table I Mutagemctty and other toxtcity data on selected aromatic ammosulphomc actds and thetr unsulphonated analogues*
IARC (1987)
Fassma (1990) Btrner (1988) lshtdate (1988) Ishtdate (1988) Yoshimi (1988) Mitchell (1988) Wangenhelm (1988) yon der Hude (1988) Hamson (1987) Zetger (1987)
Shahm (1989) Ishtdate (1988) Zetger (1987)
Z
NH 2
88--44-8
CH,NOS
NH 2
~
137-51-9
C6H?NO6S2
SO3H
S03H
98-37-3
C6HTNO4S
H O 3 S / ~ O H
HO3S
+
Zeiger (1988) ETAD (1987) NTP Rep (1986)
ETAD (1988) Shtmlzu (1983)
m- DNA m - cyt m - DNA m- GMA
c- cell trans
Yoshlml (1988) ETAD (1988) ETAD (1988) ETAD (1986)
Styles(1973)
103 CeHTNO 95-55-6
102 CTHgN 106-49-0
+ (w)
+
c+ m +/-
haembmdm8 cyt
m SAL m + (w) SAL
DNA
[contd]
Hamson (1987) Kirchner (1982)
De Flora (1984) BIBRA (1984)
Birner (1988) Neumann (1988) Pai (1985) Garner (1984) Ce-~trone (1982) c+ c+ mcm+
haembmdmg haembmdmg GMA
NTP Rep (1984) ETAD (1981)
m ÷ (w) SAL m+ SAL
fj3
~s o
f~
o
No
88-23-3
C6H6NCIO4S
N~
5857-94-3
C6H6NC104S
HO
SO3H
6370-23-6
CI ~
CI
CH 3
CsHtlNO3S
HaC ~
NH 2
Structure
SO3H
NH 3
SO3H
Zelger (1988) NTP Rep (1986)
ETAD (1987)
Ames test data Chung (1981)
Aromatic ammosulphomc aods
Table
Other data
1--contd
106 C~H6CINO 95-85-2
105 C6H6CINO 38191-33-2
No 104 CsHHN 95-68-1
+ (w)
Other data SAL SAL SAL DNA haembmdmg DNA
m + (w) SAL m + (w) SAL
m+ cc-l+ Group 3
C+
m+ m+ m+ m+
Unsulphonated analogues Ames test +
Zelger (1988) NTP Rep (1986)
Zelger (1988) NTP Rep (1985a) Shlmlzu (1983) Wdhams (1989) Blrner (1988) Yoshlml (1988) Garner (1984) DETO (1980) IARC (1987)
z
oo
12
1!
10
SO3H
6471-78-9
CsHttNO4 S
NH 2
Cl"
88-51-7
C7HsCINO3S
CH 3
1
NH 2
~
88-53-9
CTHsCINO3 S
12I-t3
Cl ~ / N H a
CH30"
~
SO3H
SO3H
CH 3
ETAD (1986)
ETAD (1986) Shtmlzu (1985)
Chung (1981)
rn DNA m DNA
m- DNA m- cyt
YoshJml (1988) ETAD (1986)
Wflhams (1989) ETAD (1988)
109 C7HsCIN 95-74-9
108 CTHsC1N 7149-75-9
107 CaHItNO 120-71-8
+ (w)
pulm ad
SAL SAL
[contd]
Ennever (1986) NTP-TR-145 (1978) DNA
m+ c
NTP Rep (1989a) Zelger (1987) Ennever (1986)
IARC (1987)
Maronpot (1986) NTP Rep 85-002 (1985) Vaimo (1985) Garner (1984) NTP-TR- 142 (1978)
Zeiger (1988) NTP Rep (1986)
m + (w) SAL mSAL mSAL
Group 2B
C+
C+
C+
C+
c-
m+ m+
~D
> >
o
0
g~
O
15
14
13
No.
5123-63-7
CIoHIsNO3 S
32432-55-6
C9H|4N203S
CH 3
-~ NH 2
/
HO3S ~-
-,~
CH3
N%
3577-63-7
CTHTNOsS
SO3H
H3C ~
HOOC
NH 2
Structure
--
-
--
ETAD (1989)
ETAD (1989)
NTP Rep (1984)
Ames test data
AromaUc ammosulphomc actds Other data
Table l--contd
112 CIoH~sN 91-66-7
I11 CgH 14N2 3102-70-3
-
-
110 C7HTNO2 118-92-3
test
No
mmm-
Group 3
c-
m m m+ m+ m-
SAL SAL DNA
SAL SAL GMA cyt DNA
Other data
Unsulphonated analogues Ames
Zelger (1988) Shlmlzu (1983) Yoshlml (1988)
IARC (1987)
Zelger (1987) NTP Rep (1984b) Mitchell (1988) NTP Rep (1985b) Cesarone (1982) NTP-TR-36 (1978)
z o-
19
18
17
16
I
-
~
H~~
N
SO3H
~
I
117-61-3
CI2HI2N206S2
N
3365-90-0
CI2HI2N206S2
3365--89-7
CI2HuN03S
98-40-8
C~I13NO3S
SO3H
~
~
S%H
NHC2H5
-
ETAD (1989)
Ashby (1982)
ETAD(1989)
¢
cell trans Ashby (1978)
115
115 CI2HI2N 92-87-5
il4 CI2HIIN 92-67-1
113 CgHI3N 94-68-8
c + Group I
c + Group I
c + Group I
IARC (1987)
IARC (1987)
IARC (1987)
[contdl
>
O
)0
O X
o
22
21
81-16-3
CloH9NO3S
SO3H
84-86-6
CIoH9NO3S
SO3H
81-11-8
C14HI4N206S2
SO3H
Structure
2N-Q-c.=c.
No.
SO3H
NH 2
-
Le (1985) Pogodma (1984) Purchase (1978)
Kler (1986) Chung (1981)
NTP Rep (1990) ETAD (1989) Zelger (1987)
Ames test data
Aromatac amlnosulphomc acads
GMA cyt
c e-
m- DNA m - cyt
c
m m
ETAD (1989) ETAD (1989) Pogodma (1984) Della Porta (1982)
D E T O (1980)
ETAD (1989) NTP Rep (1989b)
Other data
Table 1--contd
118 CIoH9N 91-59-8
117 CIoH9N 134-32-7
116 C14HI4N2 621-96-5
No
+
+
test
Ames
c+
Group 3
m-
Group 1
DNA
GMA cyt cyt cyt
mm+ mm+ C-
SAL SAL SAL SAL
m+ m+ m+ m÷
Other data
Unsulphonated analogues
IARC (1987)
IARC (1987)
Fassma (1990) lsh~date (1988) Isbldate (1988) NTP Rep (1988) Vamlo (1985) Brambdla (1985)
lshldate (1988) Zelger (1988) NTP Rep (1986) Kler (1986)
z
bO
26
25
24
23
1 17-26-4
Cl0H9NO6S 2
SO3H
SOaH
86-60-2
Ct0H9NO3S
93-00-5
C)0H9NO3S
~
HO3S~
81-05-0
C1oHgNO3 S
SO3H
NH 2
NH2
-
-
-
-
-
-
Kmr (1986) Purchase (1978) Garner (1977)
Le (1985)
Le (1985)
Le (1985)
C-
c-
cell trans
c pulm ad
c pulm ad
Purchase (1978) Purchase (1978)
Thetss (1981)
Thelss (I981)
118
118
118
118
+
+
C+
C+
C+
C+
Group 1
Group 1
Group I
Group 1
[contd]
IARC (1987)
IARC (1987)
IARC (1987)
IARC (1987)
o
o
o
29
28
27
No.
HO3S I
118-03-6
CIoH9NO9S 3
v
SO3H
~
118-33-2
Cl0H9~O6S2
SO3H
86-65-7
C10HgNO6S 2
~
H03S ~
HO3S
SO3H
Structure
~SO3H
NH 2
NH2
-
-
-
ETAD (1989)
ETAD (1989)
ETAD (1989)
Ames test data
Aromatic ammosulphonlc acids
c +~ pulm ad Theiss (1981)
Other data
Table l---contd
I 18
118
118
No
+
+
Ames test
c+
c+
¢+
Group I
Group 1
Group 1
Other data
Unsulphonated analogues
IARC(1987)
IARC (1987)
IARC (1987)
32
31
30
HOsS~
H03S
90-51-7
CIoH9NO4S
HO
5639-34-9
CtoH9NO4S
CIoH9NO4S 116-63-2
SO3H
~
/OH
OH
ETAD (1990) Freeman (1987)
Chung (1981)
ETAD(1987) Kler (1986) Chung (1981) m + cyt
Gayathn (1980)
CRoHgNO 4384-92-3
120
119
119 C~oH,NO 2834-92-6 Wdlmms (1989)
DNA
m-
[contd]
Kter (1986) Dabney (1985) Garner (1977)
SAL SAL SAL
m mm-
f~
o
P~
e~
o
H
O
HO3S ~
.NH2
3
/
~
CIoH9NO7S2
S
SO3H OH
90-20-0
CIoHgNOTS2
OH
NI-12
SO3H
SO3H
OH
-
-
-
ETAD (1989)
Freeman (1987)
Chung (1980
Ames test data
m
DNA
m + GMA m- cyt
ETAD (1990)
Palmer (1979) Palmer (1979)
Other data
Table I--contd
122 C~0HgNO 606-41-7
CIoHgNO 2834-9 I-5
12l
119
No
test
Ames
Other data
Unsulphonated analogues
+ =posmve response - = negative response w = weak response ~ = quesUonnable results c =carcmogemoty, oral apphcatlon cell trans =cell transformation study haembmdmg= haemoglobin adducts m = genotoxmlty/mutagemcltym cyt = cytogemc~ty/chromosomemutation m DNA = DNA repair/indicator test m GMA = gene mutatton assay m eucaryotm cells (not Ames/Pnval) m SAL = Salmonella/mlcrosome (Ames) test pulm ad = pulmonary adenoma carcmogemclty test *With the exception of the Salmonella/mlcrosome assay, only a selecuon of the available data is shown For human carcinogens, only the IARC classlficaUon ts menUoned IARC classification. Group 1, carcmogemc to humans, Group 2B, possibly carcmogemc to humans, Group 3, not classifiable as to its carcmogemclty to humans
35
34
2007-20-7
CIoH9NO7S2
NH 2
33
H3OS
Structure
No
Aromatic ammosulphomc aods
z
Genotoxlclty and carcmogemcity of AASAs attempted to validate this test system using 65 chemicals together with available results of 2-year carcinogenicity and in vitro mutagemcity tests. The authors concluded that there was a poor agreement between genotoxicity results and the results obtained with strain A mice. Furthermore, results obtained with strain A mice are also not in agreement with results from 2-year carcinogenicity tests on aromatic amines and several other miscellaneous chemicals. Based on these findings, the concern of Theiss et al. expressed in their previous paper (Theiss et a l , 1981) about the possible carcmogenic potential of AASAs is certainly not justified. The aim of this paper ~s to investigate whether the available toxicity data on AASAs involved in dyestuffs chemistry give indications of any genotoxic or carcinogenic potential. The role of aromatic aminosulphonic acids as dyestuff intermediates and metabolites The sulphonated derivatives of the aminonaphthalenes and anilines are particularly important dye intermediates. The sulphonic acid group is used extensively in the dye chemistry to confer water solubility and for its ability to act as a good leaving group in nucleophllic substitutions. It is used almost exclusively for these purposes since it has only a minor effect on the colour of a dye. The amino group is the single most important functional group in dye chemistry. In azo-dye intermediates, it not only provides access to the azo group (by diazotization and coupling), but is also the most widely used auxochrome as can be seen from its presence in so many coupling components. Dye intermediates and similar compounds are an important group of primary metabolites of azo dyes, which constitute over 65% of all dyestuffs. Azo dyes all share one or several azo ( - - N : N - - ) groups, but they constitute a large variety of differing structures with a wide range of physlcochemical properties: from extremely water-soluble dyes to practically insoluble azo pigments, and from anionic and cationic to nonionic dyes. It has already been shown that under anaerobic conditions a variable percentage of the azo groups can be reduced enzymatically to the corresponding sulphonated and unsulphonated aromatic amines in bacteria and in mammalian systems including humans. In bis- and trisazo compounds the metabohc reduction of the azo groups proceeds stepwise under intermediate formation of the partly reduced molecules (i.e. azo-amino compounds). The formation of carcinogenic aromatic amines is assumed to be the major causative factor of the carcinogenic response to a given dye in an animal lifetime bioassay. Typical examples are the benzidine dyes, which are metabolized to the known human carcinogen benzidine. Based on present knowledge, it might be assumed that azo dyes, which are metabolized to a substantial
647
extent to carcinogenic amines in the living organlsms, should be suspected as potential carcinogens. Conversely, if non-carcinogenic amines are formed, the probabihty that the parent compound has carclnogenic properties becomes very small. It must be emphasized that beside this reductive cleavage of the azo group, other metabolic transformations could occur in the sense of additional activation or deactivation. Genotoxicity data of aromatic aminosulphonic acids As already mentioned, AASAs constitute a large number of potential metabolites especially of anionic dyes. Although few bioassay data on carcinogenicity exist, much information is available on genotoxiclty. This paper compiles a substantial amount of published and unpublished mutagenicity data. Table 1 displays available results of the Salmonella/microsomc test (Ames test), and other genotoxiclty and carcinogenicity data for 35 AASAs. For comparison, some available data on the unsulphonated analogues are also given. In contrast with mutagcnic and carcinogenic amines, such as 4-aminobiphenyl, benzidine and 2naphthylamine, the sulphonated analogues are not mutagenic in the Ames test. In the case of 2-naphthylamine several aminosulphonic acid isomers were non-mutagenic (Le et al., 1985). In a recent publication (Freeman et al., 1987), the results from the Ames and Prival mutagenlcity test of six aminonaphthalenesulphonic acids were reported. Five of them showed a very weak positive response (i.e. 1amino-naphthalene-7-sulphonic acid, 1-amino-8-hydroxynaphthalene-4-sulphomc acid, 2-amino-6-sulphonic acid (24), 2-amino-8-hydroxynaphthalcne-6sulphonic acid 32 (g-acid) and 3-ammo-8-hydroxynaphthalene-6-sulphonlc acid (y-acid)), whereas l-amlno-8-hydroxynaphthalene-3,6-disulphonic acid (34) was negative in both tests. The authors presume that the very weak positive response exhibited by the five amlno-naphthalenesulphonlc acids might be due to impurities since the samples used in their assays were not 100% pure. Such an assumption is supported by tests with highly purified materials. Le et al. (1985) found 2-aminonaphthalene-6-sulphonic acid (24) to be non-mutagenlc in the Salmonella/ microsome test in the presence of hepatic S-9 derived from the rat, hamster, monkey and humans. In the same test, using S-9 derived from the rat, 2amlno-8-hydroxynaphthalcne-6-sulphonic acid (32) (),-acid) (ETAD, 1990) and very recently 3-amino8-hydroxynaphthalene-6-sulphonic acid (J-acid) (ETAD, 1991) gave a negative response (not listed in Table 1). Similar experiences regarding purity were encountered with 2-amino-5-methylbenzenesulphonic acid (5) (4B-acid) and 2-amino-5-chloro-4-methylbenzenesulphonic acid ( l l ) (CLT-acid), which were negative in the Ames test only after thorough
No
HO3S -
-
~
C6HTNSO3
-
Structure
NH 2
Aromattc aminosulphomcacid (hypothetical dye metabohte)
N~N
C.I. 19140 C.I. Acid Yellow 23
NaO3S---~
C.l. Food Yellow 3
C.I. 15985
NaO3S~
C.I, 14600 C.I. Acid Orange 20
SO3Na
COONa
HO ~-----N~SO3Na
HO
Borzelleca (1988a) Borzelleca (1988b) Maekawa (1987)
IARC (1987) NTP-TR-208 (1981)
BIBRA (1988)
Azo dye wRh negaUveresults m caremogemcRyassays IARC (1987) Schach von WIttenau (1983)
Table 2. Ammes from azo dyes with negatwe results m caranogematy assays*
I01
No.
Unsulphonated analogue of amine
l0
CH30
/~.~-"-
. CH3
CaHItNSO 4
HO3S
T
NH~
CoHTNSO3
H03S
NH2
HO
N= N
C.I. 16035 C.I. Food Red 17
NaO3SH 3 C ~
cH3
OCH 3
C.I. Food Red 1
C.L 14700
NaO3S
C.I. 13065 C.I. Acid Yellow 36
NaO3S~__
C.I. 15980 C.I. Food Orange 2
Na03S
HO
NaO3S
HO
SO3Na
NH©
SO3Na
Borzelleca(1989)
IARC (1987) Schach von Wittenau (1983)
Vettoraz~ (1981) ADL (1980)
Vettorazzl (1981)
[contdl
107
101
> >
o
=s
o
21
No
C10H9NSO3
SO3H
NH 2
Structure
AromaUc amlnosulphomcacid (hypotheUcal dye metabohte)
C.I. 20285 C.I. Food Brown 3
NaO3S
N~N
SO3Na
OH
N~N
HO
N~N
C.I. 14720 C.I. Acid Red 14
C.I. 16185 C.I. Acid Red27
NaO3S~
SO3Na
N~-----NS
~
SO3Na
SO3Na
C~OH
HO
HO
SO3Na
Vettorazzi (1981) Drake (1978) Carpamm (1978)
BIBRA (1990) NTP-TR-220 (1982)
ADL (1980) Clode (1987)
Azo dye with negative results m carcmogenlcRyassays
Table 2 - - - c o n t d
107
No
amine
Unsulphonated analogue of
o~
37
36
CsHIINSO3
c~
~ C ~ s 0 3
NH2
CsHIINSO3
CH3
C H 3 ~
NI-I2
H
SO3H
NaOsS
i~c
NaO3S
CH3
N
O3Na ~
C.I. 14700 C.I. Food Red 1
c~
HO
~
NaO3S
N
HO
C.I. 16255 C.I. Acid Red 18
C.I. 14815 C.I. Food Red 2
NaO3S
SO3Na
HO
SO3Na
IARC(1987) SchachyonWRtenau(1983)
Vettorazzl(1981)
Brantom(1988) BIBRA(1986) Mason(1974)
[conta]
123
123
~
r.~
0
g~ e~
o
g
38
!1
NO.
HO3S
HOsS
HOsS
"CH3
C IoHIoN2SO3
NH 2
CTI'IgNSOs
CHs
CTI~NSO3CI
Cl
N~
Structure
Aromattc amlnosulphonicacid (hypothetical dye metabohte)
N
HO
NaO3S---O
C.I. Food Black 1
SO3Na
COONa
NaO3S
HO
SO3Na
NHCOCH 3
BIBRA (1989b) Vettorazzl (1981) Drake (1977) Gaunt (1972)
BIBRA(1982a)
BIBRA (1989a) IARC (1987) NTP-TR-225 (1982)
Azo dye with negattve results in carcinogemcttyassays
C.I. 28440
C.I. 15850 C.I. Pigment Red 57
N
O3Na
C.I. Pigment Red 53:1
C.I. 15585:1
N
SO3Na
CHs--~
CI
OH
Table 2---contd
124
102
No 108
2nllne
Unsulphonated analogue of
Z
CIoH9NSO4
H O 3 S / ~
NH2
OH
t
C.I. 16035 C.I. FoodRed 17
H3C
C.I. 15985 C.I. FoodYellow3
NaO3S ~N'---N ~
HO
N_~_NHO~
C.I. 15980 C.I. FoodOrange2
NaO3S~
SO3Na
SO3Na
SO3Na
Borzelleca (1989)
BIBRA (1988) IARC(1987) NTP-TR-208(1981)
Vettorazzt(1981)
[cored]
!19
r~
>.
o
o
33
No. 39
HO3S
CI0HgNS207
NH2
CIoHgNS207
HO3S ~ O H
Structure H03S
Aromaticammosulphomcacid (hypotheucaldye metabohte)
SO3H
NaO3S
NaO3S - - ~
N = . N - ~
C.I. 16255 C.I. Acid Red 18
N~ N
C.I. 16185 CT Acid Red 27
N
N
SO3Na
S03Na
SO3Na
HO SO3Na
C.I. 16230 C.I. Food Orange 4
Na03S
HO
IARC (1987) Clode (I987)
Zeiger (1987) NTP-TR-211 (1987) IARC (1987)
Brantom (1987) BIBRA (1986) Mason (1974)
Azo dye with negativeresults m carcinogenicttyassays
Table 2 - - c o n t d
119
119
No.
Unsulphonated analogue of amine
o~
41
40
j,.~ NH2
/NH 2
CIoH9NSO4
SO3H
~
OH
C toH9NS04
SO3H
OH
NaO3S
N
C.I. 14700 C.I. Food Red 1
C.I. 14815 C.I. Food Red 2
• SO3Na
HO
N
HO
NaO3S
C.I. 14720 C.I. Acid Red 14
SO3Na
OH ~
SO3Na
SO3Na
Vettorazzl (1981)
Schachyon Wittenau(1983)
IARC (1987)
BIBRA(1990) NTP-TR-220(1982)
[contd]
119
119
u.
o'J
o
o
o
HO3S
SO2I-IN. OH
OH
CloH1oN2S207
H2N
C12HI4N2S20s
SOsH
CITHI6N209S 3
H O s S / ~
H,C----~
Structure
So3NH2H
©
©
~
~
OH
NH2
C.I. 17200 C.I. Acid Red 33
NaOsS
N~N
N
SOsNa
SO3Na
NHCOCH3
C.I. 18050 C.I. Food Red 10
NaO3S
N
HO
~
SO3Na
_ NHO2S
C.I. Acid Red 106
C.I. 18110
NaOsS
N~-N ~
HO
~ CI-I~
BIBRA(1982b) BIBRA(1982c)
VettorazzJ(1981) WHO(1977)
IARC/WHO(1981)
Azo dye withnegativeresultsin carcmogemcltyassays
127
126
125
No
amine
Unsulphonated analogueof
*ForaromaUcammosulphomcacids withnumbers 1-35and for unsulphonatedanalogueswithnumbers101-122,see Table 1 (mutagemcltyand othertoxicitydata of selectedaromaUcammosulfomcacidsand their unsulphonatcdanalogues).
44
43
42
No.
Aromattcammosuiphomcacid (hypotbeucaldye metabohte)
Table 2--contd
Z e~
657
Genotox~clty and carcmogeniclty of AASAs purification. Both substances were also negative in the micronucleus assay (ETAD, 1988). The two compounds reported by Freeman et al. (1987) to be weakly positive m the Ames test (i.e. lamino-naphthalene-7-sulphonic acid and 1-amino-8hydroxynaphthalene-4-sulphonic acid) are not listed as mutagemc in Table 1 of this review in view of the uncertainty regarding the purity of the samples tested. Unlike benzidme (115) and 2-aminonaphthalene (118), aniline (101) and p-ammotoluene (102) have only weak (aniline) or no significant carcinogenic potential m animal studies and are not mutagemc in the Ames test. The sulphonated analogues of aniline and p-aminotoluene are therefore, as expected, not mutagenic in the Ames test. In some cases, methyl and chlorine subst~tuents m the amline molecule considerably influence the genotoxic or carcmogemc potency. Several of these chemicals are mutagenic m the Ames test. The sulphonated derivatwes, however, as shown in Table 1, were not mutagenic m the Ames test. In addition to the results from the Ames test, some of the compounds gwen in Table 1 have been studied in other tests on genotoxiclty (m vivo and m vitro) including cell transformation assays. The outcome of all these studies reflects, with only few exceptions, the absence of genotoxicity for AASAs. 2-Ammonaphthalene-1,5-disulphonic acid (26) and the disulphonlc acid derivatives of benzldine, (18) and (19), were not active in cell transformation assays m vitro, whereas the unsulphonated carclnogemc amines induced cell transformation m vitro In contrast to 2-aminonaphthalene (118), which has been reported to cause D N A damage (positwe m several UDS tests), the l-sulphonic acid analogue (22) was negatwe in th~s test system. 2-Aminonaphthalene induced gene conversion in Saccharomyces cerewsiae with and without metabolic activation. 2-Naphthylamine-l-sulphonic acid (22) did not produce gene conversion m trp or ade genes of S. cerevistae strata D4 (Della Porta et al., 1982). Supportive evidence of non-genotoxlclty for AASAs also comes from mutagenicity studies with azo dyes usmg reductive assay conditions. Structures of Direct Black 19 analogues with unsulphonated p-phenyleneblsazo moieties were active m the Salmonella/microsome assay, whereas the sulphonated derivatives were not (Lm and Solodar, 1988). To summarize the results, in general aromatic phenyl- and naphthyl-aminosulphonic acids are nonmutagenic to Salmonella. Other tests on genotoxiclty have also been considered. The data compiled show no or very low mutagenic potentml. In addition, three AASAs were negative in feeding studies on carcmogenicity. The available data indicate that sulphonation of phenyl- and naphthylamines decreases toxicity considerably. Where comparable data on sulphonated and unsulphonated analogues exist, loss
of genotoxlc and/or carcinogenic suiphonation is observed.
activity
on
Mechanistic considerations It has been repeatedly reported (Combes and Haveland-Smith, 1982; Garner and Nutman, 1977) that sulphonat~on renders amines less mutagenic and non-carcinogemc, and azo dyes based on AASAs non-carcinogenic (see Table 2). It appears to be a general rule that highly sulphonated azo dyes are poorly absorbed from the gut (Walker, 1970), and therefore become b~oavailable to a significant extent only after azo reducuon to the corresponding ammosulphonic acids. The degree of absorption from the gut is dependent on the polarity of the molecule, and sulphonation increases the polarity of the molecule For sulphonated amines, significant absorption has been observed (for review see Walker, 1970). Monosulphonated amines (e.g. sulphanilic acid, naphthlomc acid) are absorbed to a large extent, possibly m the zwitterionic form. Unlike aromatic amines, however, ring substitution by sulphonation of azo dyes based on AASAs prevents the activation of amino groups by liver metabolism (Garner and Nutman, 1977). Lin and Solodar (1988) beheve that sulphonation (as in the case of p-phenylenediamine) Increases the electronegatlvity of the metabolized molecule and thus decreases the electrophilicity required to attack D N A bases. Recently, Levme (1991) m h~s review on metabohsm of azo dyes explained that highly charged sulphonated azo dyes resist bacterial enzymic attack, and are poorly absorbed from the intestinal tract thus providing poor access to the hver endogenous metabolism As m the case of the non-carcmogemc, non-mutagenic 2-naphthylammo-l-sulphomc acid, pharmacokmetic studies indicate a rapid elimination of unchanged aminosulphonic acid after oral or iv administration. No biotransformation or cleavage of the sulphonic group was detected (Della Porta, 1982). As a more general conclusion to a study on the structural bas~s of the mutagemc~ty of phenylazoanihne dyes, Rosenkranz and Klopman (1989) reported that sulphonation appears to interfere with a structure in the azo dyes that is required for oxidation of the amino group A negatwe inductive effect of the sulphomc group may influence azo reduction or amine oxidation Combes and Haveland-Smlth (1982) suggested that the sulphonlc group may exert an unfavourable electromc influence on the activation of the 4-amino-Natom by exerting a strong negative inductive effect. The negative inductive effect of the sulphonic acid group on azo reduction has been demonstrated; Walker and Ryan (1971) showed that analogues of Red 2G and Red 10B sulphonated in the p-position of the phenyl ring were reduced by cell-free extracts
658
R. JUNG et al.
of Streptococcus faecahs two to three times more rapidly than the parent compound, owing to the substitution with an electron-withdrawing group.
Absence of evidence of the carcinogenic potential of aromatic aminosulphonic acids Few carcinogenicity studies have been carried out on substituted amlnonaphthalene- and aminobenzenesulphonic acids, probably because these compounds are expected to be non-carcinogenic. However, the existence of a considerable number of negative bioassays of azo dyes, which are metabolized to such aods, give additional evidence of their low chronic toxicity and their non-carcinogenic (or at most low carcinogenic) potential. It has been demonstrated that azo dyes are metabolized in the organism to aromatic amines by reductive cleavage of the azo group in bacterial systems both in animals and humans. However, the rate of this reaction is very much dependent on the individual structure and can differ widely. This has been shown by Watabe et al. (1980) in their study on the reduction of four azo dyes by micro-organisms from human faeces. The formation of AASA metabolites is certainly not always quantitative. There are several factors influencing the rate of formation such as other competing metabolic processes, rate of absorption, route of admimstratlon, low capacity of the reductlve enzymes, lack of availability of the dye to these enzymes, steric hindrance, etc. Table 2 lists azo dyes for which adequate negative 2-year bioassays have been done. With a few exceptions, the dyes can hypothetically form only AASAs (also compiled in Table 2) on reductive cleavage of the azo group. Considering the very high doses applied in most of the studies shown in Table 2, it can be assumed that substantial quantities of the corresponding AASAs have been formed in the gastro-intestinal tract and/or in the liver. The lack of tumorigenic response in these studies is indicative of a very low or even absent carcinogenic potential of these metabolltes. This conclusion is supported by the genotoxicity data summarized in Table 1. It is generally accepted that genotoxicity is a primary event in the carcinogenic process. Therefore the absence of genotoxiclty with amlnosulphonic acid supports the conclusion that these compounds do not act as genotoxlc carcinogens. Table 1 also includes negative carcmogeniclty bioassay data for several aminosulphonic acids. In the case of benzidine and 2-naphthylamine, it has been shown experimentally that sulphonation of the parent carcinogenic amine leads to non-carcinogenic substances. Although there is a long history of production of a large number of AASAs in the dyestuff manufacturing Industry, there are no indications of any carcinogenic risks from these compounds. In
contrast with these findings, bladder cancer cases resulting from exposure to 2-naphthylamine and benzidine are well known. An interesting observation in this connection was made by Kumar et aL (1981), who found in their immunological studies a correlation between increased lymphocyte reactivity and bladder cancer risk in workers exposed to 2-naphthylamme (118). Workers exposed to 2-naphthylamine-l-sulphonic acid (22), but not to 2-naphthylamine, had lymphocytes with a normal range of reactivity, which is in keeping with the suggestion that this compound is non-carcinogenic.
Conclusion The evaluation of available information and data on AASAs, either in use for dyestuff manufacture or being formed by reductlve cleavage of azo dyes, shows that these compounds, unlike some known carcinogenic aromatic amines, generally have no or very low genotoxic effect and hence no or minimal tumorigenic potential. In addition, considering the low exposure to AASAs prevailing in the techmcal application and use of azo dyes, there is not likely to be any significant toxic risk if AASAs are formed by metabolic cleavage in the organism or in the environment, or occur as micro-contaminants in colourmgs.
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659
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660
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