Review of the genotoxicity of formaldehyde

Mutation Research, 196 (1988) 37-59

37

Elsevier

MTR07255

Review of the genotoxicity of formaldehyde Te-Hsiu Ma and Mary M. Harris Institute for Environmental Management and Department of Biological Sciences, Western Illinois University Macomb, IL 61455 (U.S.A.) (Received 20 July 1987) (Revision received 4 February 1988) (Accepted 15 February 1988)

Keywords: Formaldehyde, genotoxicity, review

Contents Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Metabolism of formaldehyde . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Genotoxicity of formaldehyde . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . I. Mutagenicity . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . II. D N A damage, repair and inhibition of synthesis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . III. Clastogenicity - - Chromosome damage and sister-chromatid exchange . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . IV. Cell transformation and carcinogenicity . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . V. General toxicology and teratogenic effects . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . VI. Genotoxicity of related aldehydes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . VII. Epidemiological studies and risk assessment . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Conclusions and discussion . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Acknowledgements . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Introduction

In our so-called civilized society, formaldehyde (FA) is the most prevalent man-made pollutant in the air. Its ubiquitous existence in and out of our living and working places poses a serious threat to human health as well as to the well-being of other living organisms. The annual production of formaldehyde for industrial, agricultural and other processing purposes by United States alone was around 6.4 billion pounds in 1978 [17] and 7 billion pounds in 1982 [81]. The annual production was around 9 billion pounds [96] in 1983 and

Correspondence: Dr. Te-Hsiu Ma, Ph.D., Institute for Environmental M a n a g e m e n t , W e s t e r n Illinois University, Macomb, IL 61455 (U.S.A.).

37 39 40 40 42 45 47 48 49 49 50 52 52

5 billion 860 million pounds in 1986. It has been utilized by numerous kinds of industries and professions, and more than a dozen names have been applied to this single compound, such as formalin, formic aldehyde, formal, fyde, ivalon, karsan, lysoform, methylene oxide, morbicid, oxomethane, oxymethylene and paraform [17]. Some of these agents remain in liquid or solid forms and the fumes are constantly released into the air through evaporation and offgassing from man-made objects. The major sources of FA fumes could be roughly categorized into three groups: those generated from industry, those released in residential housing, and those fumes in various occupational indoor settings. A large quantity of the agent was used in the form of FA-based resin for binding of plywood, particle board, dyes, and textile fibers

0165-1110/88/$03.50 © 1988 Elsevier Science Publishers B.V. (Biomedical Division)

38 [4,144]. Other industries such as furniture construction, leather tanning, paint and lubrication, upholstery, carpeting, drapery and garments [4], and common household commodities such as cosmetics, pesticides, and disinfectants are directly or indirectly involved with FA. The fumes are released in the household through the process of offgassing of fabrics, wall paneling, carpet, paper plates and cups [157]. The other indoor sources are from combustion of coal, oil, kerosine, tobacco smoke [203] and auto exhaust fumes. In this group, mobile homes which house 5% of the U.S. population [81] are unique structures being extraordinarily rich in formaldehyde fumes [70,113,114]. An extra class in the indoor category is the houses insulated with urea-formaldehyde foam [197,198]. This practice turned the old houses into a new FA fume trap. It has created serious health problems [106] across the country. Although such practices were banned by U.S. EPA in 1970s the problems of the FA fumes in the insulated houses linger on. The third group is the occupational indoor pollutants, such as embalming fluid [203], specimen and food preservatives, paraformaldehyde paste and formocresol used in dentistry [112], and disinfectants in hospitals and laboratories. Since the human health effects of FA are the major concerns in our scientific communities, numerous studies have been carried out on the immediate effects of the exposure. Most of these were conducted through questionnaires and interviews and the results were mixed with objective factual statements and subjective opinions. The minimum concentration of FA fumes which is detectable by the olfactory sense and at the same time sensed by eye irritation is around 0.1-1.0 ppm [14,115]. The mild case of ill-effects can be created by 1-5 ppm concentration in a 6-h duration. These include: headache, diTyiness, wheezing, runny nose, drowsiness, malaise [5], coughing, nausea in adults, while infants suffered vomitting, constant crying, diarrhea, and coughing [206]. The severe cases include sore throat, burning eye, skin irritation and rashes, conjunctivitis [144], coryza, dyspnea, insomnia, memory lapses, and asthma [197]. Of course, the degree of the reaction was often determined by the specific sensitivity and allergic tendencies of the individual [86]. Ingestion of FA [141] in large quantities results in loss of

consciousness, vascular collapse, pneumonia and hemorrhage. The long-range effects in the human population were studied through the epidemiological approach on the carcinogenicity and teratogenicity of the chronic exposure to the fumes in various occupational groups. There were indications of an elevated rate of morbidity and mortality through occupational exposure which was mixed with other factors. According to Hileman [81] there was no clear cut national policy over the control of production and application of this agent in 1982. There is no international effort to formulate a workable policy for the safe usage of this agent so far. Among the dozen of industrial nations in the world, the U.S. has the highest occupational dose standard of 3 ppm while other nations ranged around 1-2 ppm [81]. Extensive studies and evaluations were cartied out at the Chemical Industry Institute of Toxicology (CIIT) and the Institute of Environmental Mutagens of New York University (NYU) since the late 1970s. Several reviews and workshops were conducted to determine its carcinogenicity. Not until recently, the U.S. EPA claimed that FA is a possible carcinogen in humans. The regulatory action and national and international policies presumably is in the making seven years after the initial claim on its carcinogenicity [130]. While the physical and chemical technologies are advancing at a rapid rate, the methods for detection of trace amounts of formaldehyde are numerous. The most frequently used are: colometric tube [168], chromotropic acid [144] spectrophotometry, fluorometry, high pressure liquid chromatography [177] ion chromatography, polarography, gas chromatography, pararosamine and toluene extraction, and sodium bisulfite absorption [177,184]. Since the accuracy and reliability of the modern analytical instruments are very high, the existing quantity of the agent under given conditions can be measured with great precision. However, the biological effects of the agent under the different environmental conditions are not necessarily clear. Variation of environmental factors plus the possible synergistic effects of combination with other agents, may result in unexpected extraordinary ill-effects. Among the 209 papers reviewed (1976 through 1986), few mentioned the necessity of biological monitoring [197]. So far, no

39 bioassay has been applied to determine the biological effects although a number of highly sensitive bioassays could be effectively applied to give the first warning signals [116,119,120,122]. One review paper published in 1981 [17] dealt heavily on product usage and occupational safety. The report by the Federal Review Panel on Formaldehyde [169] focused on epidemiology and metabolic activities with limited accounts on mutagenicity and carcinogenicity, plus the risk assessments predominantly concerned about human health. The Report of the Consensus Workshop on Formaldehyde [168] emphasized mainly analytical methods, source of FA pollution and the site of epidemiological sampling. Current review will concentrate on the genotoxicity of this agent. There will be 7 major categories including: I. Mutagenicity; II. DNA damage/repair and inhibition of synthesis; III. Clastogenicity; IV. Cell transformation and carcinogenicity; V. General toxicity and teratogenic effects; VI. Genotoxicity of related aldehydes; and VII. Epidemiology and risk assessments. These experimental data derived from in vitro and in vivo studies of prokaryotes, plant and animals not only throw some light about the harmful effects of this agent to humans but also give the evidence of its deleterious effects on living beings of all forms. The total ecological impact of this omnipresent agent in the modern world should carry much more weight than its effect on humans alone.

Metabolism of formaldehyde The basic chemistry and chemical reactions of FA and its related molecules have been extensively elaborated in the earlier review conducted in 1977 [11]. Chemical reactions between FA and the essential biological macromolecules were also discussed [11]. The additional metabolites of FA and FA generating endogenous or exogenous compounds in mammalian systems have been scrutinized in the last 10 years. The present review will concentrate on the new additions which may lead to the better understanding of the genotoxicity of this agent. In the living organisms, FA is generally produced through the catabolic reaction of methanol or methylamine with the help of a dehydrogenase

with the exception of some microorganisms [126]. A number of endogenous and exogenous compounds of the experimental animals and humans release FA through catabolic reactions in their body. These include N-methyl-N-(hydroxymethyl) thiourea [92] which yields FA and N-methylthiourea choline, and dimethylaminoethanol, dimethylglycine, methionine [132], dimethylnitrosamine, dinitrosopentamethylenetetramine, hexamethylenetetramine. Hexamethylphosphoramide and methylene dimethanesulfonate [13] could also yield FA through biochemical reactions. Some of the foreign compounds could generate FA through the xenobiotic reactions, such as cytochrome and methanol, as well as through dehalogenation of methyl chloride. In biosynthetic reactions, FA is utilized as the intermediate C 1 units to form biomolecules through 3 different pathways [10]. In these syntheses [132], methylene tetrahydrofolic acid (methylene-THF) serves as the carrier of the C 1 units from FA into (1) serine, or (2) ribulose 5-phosphate or (3) dihydroxyacetone pathways. Methylene-THF is commonly referred to as the "Active Formaldehyde" [132] which plays the essential role in the synthesis of purine, pyrimidine, methionine and glycine [190]. Among the common metabolites, N-(hydroxymethyl)urea, N,N'-bis(hydroxymethyl) urea, thiazolidine-4-carboxylic acid, dimethylnitrosamine [6], are excreted in the urine after the injection of labelled FA in experimental animals [6,78,79,80,132]. One of the major findings in the pharmacokinetic studies using inhalation experiments with rats and mice was that FA reacts with the first site of its encounter which is the nasal mucociliary apparatus. Most of the FA fumes are absorbed in the nasal passage, little FA reached the trachea [40] thus reducing the chance of damaging the further portions of the respiratory tract. Based upon the studies of Heck et al. [76], using 14C labeled FA to determine the rate of incorporation of FA to various tissues and especially to the plasma and other blood components, a large portion of the inhaled FA is incorporated quickly into the erythroblasts with lesser amount in the plasma. Similar results were found when FA is administered orally to experimental animals. FA was incorporated into erythrocytes 5 time more than was

40 incorporated in the plasma [115]. Generally the inhaled FA is usually removed by the respiratory tract. About 40% of the oral administered FA was exhaled as carbon dioxide within 12 h [115]. Extensive studies using ~ac-labeled FA fumes to fumigate Fisher 344 rats were performed and a parallel study was carried out by bolus intravenous injection via the tail vein. The results of the inhalation treatment shows that most of the radioactivity measured by scintillation counting of the tissue samples was from the nasal mucosa, only a small amount entered the trachea and a very small amount was found in the plasma [75,76]. The pharmacokinetic studies using the intravenous injection approach gave the similar pattern of incorporation in the blood components as in the inhalation approach in a time span of 200 h. The labeled FA is mostly found in the plasma during the first few hours. The red blood cells incorporated more of the labeled FA 25 h after the treatment and remained at the high level thereafter while the labeled FA decline in the plasma moiety. The results of topical exposure of FA liquid to rats, Guinea pigs and monkeys [91] indicated that the liver and lungs had relatively higher incorporation rate and the major portion was exhaled as CO 2 since the common path of FA metabolism is by the formation of formic acid. The formic acid in turn breaks down into carbon dioxide and water [141]. The excreted portion of labeled FA was in urine, and only a small amount was found in the feces. In the studies of the effect of FA on grasshopper embryonic neuroblasts [162], it was discovered that fetal calf serum added to the tissue culture medium reduced the clastogenicity of the FA. The reduction was attributed to the high reactivity of FA with calf serum. The free FA is highly reactive with amines, thiols, hydroxyls, amides and cysteine to form adducts. Adducts of nucleophiles may give rise to hydroxymethylene compounds. Studies on DNA adducts of FA have been reported [165,166] using the 32p-labeled D N A to trace the adducts to DNA molecules. Results indicate that 5-methylcytosine is the general site of binding. It was found that FA binds readily with guanosine [77]. The adducts of FA to cysteine results in thiazolidine-4-carboxylic acid found in the urine of FA-treated rats [78-80]. The F A a d d u c t s to amines which result in

aminomethylol derivatives was claimed to have a damaging effect on D N A templates, thus influencing the process of transcription [163]. Formaldehyde was well known to be a potent agent for induction of cross-links between DNA molecules, between D N A - p r o t e i n , and between proteins [25]. In the inhalation studies in rats and mice [191] it was found that D N A crosslinks and D N A - p r o tein crosslinks were made by covalent bonds with FA. This binding processes was usually preceeded by the breakage of hydrogen bonds of the DNA molecules [141]. The binding site of DNA or RNA with FA is usually at the amino groups of the nucleotides. In the excision-repair studies, Fornace, [48-50] using alkaline elution technique demonstrated the crosslinks between DNA molecules. The same kind of linkage was found in mouse L1210 leukemia cell lines [172,173]. Using the D N A - c e l l binding (DCB) assay, Kubinski et al. [103-105] demonstrated that FA promoted the binding of D N A with the proteinacous components of the cell. Polacow [160] treated calf-thymus chromatin with 2% FA plus DNA (T A rich) and created crosslinks between histone and DNA. He claimed that the mechanism was due to the loss of the positive charges of lysine and arginine. The antitumor agent, methyl dimethanesulfonate (MDMS) was found to be able to induce crosslinks of D N A because it could generate FA through hydrolysis [13]. Ohba [153] in his studies of FA reaction with calf-thymus nucleohistone f o u n d the f r e q u e n t crosslinkage between nucleohistones and nucleohistone with DNA. The proposed reactions are as follows: R-NH 2 + HCHO ~ R-NH-CH2OH, R-NH-CH2OH + RNH 2 R-NH-CH2-NH-R'

+ H20

The binding sites are the amino groups of the protein structure [23].

Genotoxicity of formaldehyde 1. Mutagenicity (A) Point mutations During the last decade, many kinds of bioassays were developed and a number of them were

41 efficient and reliable for detection of point mutations in prokaryotes and eukaryotes. The FA-induced mutations will be discussed under 6 different categories established on the basis of organisms used for the mutation detection. They are bacteria, fungi, Drosophila, nematode, mouse lymphoma and human lymphoblasts. Most studies in this category used the liquid form of FA for treatment except one which studied FA fumes from particle boards [56] out of 33 papers published between 1977 and 1987.

(1) Bacterial systems. Among the bacterial testing systems, two species were often used, namely Salmonella typhimurium and Escherichia coli. Many different genetic stocks of these bacterial species were utilized and some E. coli strains used for testing contained plasmids from Salmonella. ( a) Salmonella typhimurium: The standard reversion tests (Ames test) were usually used in these studies. The mutant strains utiliTed by various authors were TA97, TA98, TA100, TA102, TA104, TA1535, TA1537, TA1538, TA1539, UTH8413, and UTH8414 for reversion tests. For forward mutation of trifluorothymidine, BA9, BA13 and TM677 strains were utilized. Out of 20 some studies published, the mutagenicity of FA in the reverse mutation system was about 75% for the positive [28,29,32,34,56,73,109,128,134, 161,179,194] and about 25% for negative [33,35,57,128,161,178] results. In the forward mutation tests, most of the results were positive [32,39,61,174,178,195] unless $9 activation was employed. This was also true in the reversion tests. The reduction of mutation rate by $9 was attributed to the detoxification effect on FA from the added $9 microsomes. In general, negative responses were obtained when the FA was added to the plate culture, while the preincubation approach was more efficient in the induction of mutations [179]. So far the only bacterial test that utilized FA in the gaseous state was done in conjunction with several other gaseous components. This was done on the effects of gaseous fumes released from particle board on the plated cultures [56]. In this setting, FA was the predominant gas released along with some other volatiles, including benzene, styrene, n-undecane, and tetra-

decene during the earlier part of the treatment. The mixture gave the higher revertant rate from the newly prepared board samples under smaller doses, and the revertant rate dropped when the doses were increased due to toxicity from an overdose. The FA contents reduce with time (21 days after the initial treatment) while another group of gases, including a-pinene, n-butanol, camphene, nonanal and limonene increased by time. (b) Escherichia coli: The diluted liquid FA was utilized to treat E. coli cultures in 3 separate investigations. Kosako [100] used WP2, uvrA strain (with p K M 101 plasmid from Salmonella). and streptomycin-resistant strain (forward mutation) to test the mutagenicity of FA. Positive results were obtained. The reversion test with E. coli strain B(arg - ) and B / r ( t r p - ) by Takahashi [1941 also obtained positive results. Oda [151] conducted E. coli operon B-galactosidase inactivation experiments with FA as the treating agent. A dose-related response was obtained using the enzyme inhibition as the end point. Similarly, FA induced inhibition of transcription in E. coli was also demonstrated [163].

(2) Fungal tests. Two well established assays were utilized, namely yeast (Saccharomyces cerevisiae) and bread mold (Neurospora crassa). Liquid FA was applied to the culture media of both of these fungal tests. (a) Saccharomyces cerevisiae mutation assay: A series of metabolically deficient mutant strains (his, ade, leu, lys, met, arg, trp, iso) and radiosensitive strains (rad6-1, rad3-12) were used for reversion tests, and positive responses were obtained [241. (b) Neurospora crassa: The adenine-requiring strains ad-3A, ad-3B (38701), and the excision-repair deficient strain (uvs-2) were utilized for determining reversion frequencies induced by FA in liquid form. The two-component heterokaryons for ad-3A and ad-3B (closely linked) was constructed for tests of point mutation as well as for the multilocus deletions. Among a list of 11 wellknown mutagens tested [37,38], the mutation rate induced by FA was the lowest (3/107). The conidia of the uvs-2 excision-repair-deficient [211 ad-3 strain heterokaryon H12 and H59 were treated with FA for the forward mutation, multilocus

42 deletion and intracistronic mutation. H59 was more sensitive to FA than H12, and FA was declared to be a potent mutagen according to the response of H59 strain under this test system.

(3) Nematode (Caenorhabditis elegans). The unc-22 region of the linkage group IV was treated with FA solution to determine the forward mutation rate. The 3 × 10 -5 and 2 x 10 -4 mutation rates were induced by 0.1% and 0.07% FA concentrations respectively. Higher concentrations ( > 0.1%) were lethal and lower concentrations ( < 0.01) were ineffective [136]. (4) Drosophila melanogaster. This investigation was not designed to test the mutagenicity of FA in a conventional sense but rather using FA as the mutagen to probe the base-pair changes in the exon and introns of the known mutant genes [158]. Four A d h - mutants, Adh f"4, Adh fn6, Adh fn23 and Adh f"24 were induced by FA [15]. These mutant genes were sequenced for their base-pair deletions. Results indicated that there were 2 deletions within the intron and 2 deletions within the exon with the loss of 6 to 34 base pairs. This new approach on the studies of the molecular change of the mutant genes will be essential for the better understanding of the mechanisms of mutagenicity of FA. (5) Mouse lymphoma TK mutation. Positive results were obtained in T K locus mutation when treated with FA (0.3-0.5 g l / m l ) in mouse lymphoma (L5178) cell line [22]. (6) Human lymphoblasts. The TK6 cell line was utilized to test the mutagenicity of FA [62]. The effective dosage utilized was 130 ~tM FA concentration, in a 2-h treatment. Results of this study showed an increased rate of trifluorothymidine-resistant mutants in the FA-treated cell cultures of human lymphoblasts. (B) Recessive and dominant lethals and somatic mutations Most of the lethal mutation studies were conducted with Drosophila or mouse. The recessive lethals were usually induced in the sex-linked traits in Drosophila. Dominant lethals were presumably

the results of chromosome deletions, missing chromosomes or other drastic damage in germ cells of the treated organisms.

(1) Recessive lethals in Drosophila. Nasrat et al. [138] treated larvae of OR-K strains with 0.14% FA in conjunction with the variation of temperature (15 ° , 25 ° and 3 0 ° C ) and looked for the sex-linked recessive mutations in 3 broods of flies. Major findings in this study were that low (15 ° C) or high (30 o C) temperature treated flies had low fertility, and 25 ° C treated group had the highest rate of mutation (2/425). Woodruff et al. [207] treated mature files by feeding 1.2% FA and by injection with 0.2% FA for induction of sex-linked recessive lethals and reciprocal translocations. A 0.38% mutation rate was obtained from the injection treatment but no effect was shown by the feeding approach. (2) Dominant lethal in mouse. The Q strain of mice was utilized in two consecutive studies for dominant lethal [46,471. A 50 m g / k g i.p. injection gave embryonic death or pre-post implantation death at 1 week and 3 week periods. The combination of FA (30 m g / k g ) and hydrogen peroxide (90 mg/kg) treatment, showed positive results in the pre-implantation death, and a slight increase of chromosome fragments in animals treated with these mixtures. (3) Somatic mutation. Szabad [193] studied sex-inked recessive lethal and mosaicism in eye cells resulting possibly from nondisjunction, mitotic recombination, chromosome breaks, missing chromosomes or somatic mutation. The FA liquid dosage was 50 mM, and was applied to the larval medium. The FA effect was compared with 1 kR of X-rays. For the induction of mosaics, FA was less efficient than the 1 kR of X-rays. For induction of recessive lethal, FA was more efficient than 1 kR of X-rays. Treatment to the adult flies showed no effect at all. IL DNA damage, repair and inhibition of synthesis Owing to the advances made in the biochemical and cytological techniques in the last few decades, detection and measurement of DNA alteration, repair and inhibition of synthetic activities can be

43 achieved quickly with a high degree of accuracy. Extensive studies of the FA effects on human bronchial cells or fibroblasts were carried out. In addition, specialized neoplastic human tissues as well as HeLa cells were also utilized for induction of DNA damage. Besides human cells, monkey kidney, Chinese hamster ovary (CHO), rat tracheal epithelium, mouse leukemia L1210 cell line and Drosophila larva in the animal kingdom were also utilized. Although many kinds of cell cultures of higher plants were available for DNA studies, only yeast and Escherichia coli of the relatively lower forms were utilized according to the current collection of literature. A general discussion will be given here under the category of organisms used.

(1) Escherichia coll. Strain CM871 is a carrier of triple mutants, LexA, recA and uvrA [199]. The induced D N A damage was effectively expressed in the reduced survival rates. Results of this study indicated that there was a drastic reduction of survival rate (0.5%) at the 7.8 # g / m l dosage. Mitsevich et al. [135] treated the wild-type E. coli (K-12 or B) and the repair-deficient strains with FA at 10 -2 , 5 × 10 -3 , 2.5 × 10 -3 , and 1.25 -3 M concentrations and showed that the survival rate was inversely proportional to the dosages. (2) Saccharomyces cerevisiae. N 123 and 2 UV-sensitive mutants, radl_ 3 and rad3_ 5 and RAD (wild) were used [123-125]. Single-stranded breaks (SSB) were induced by FA at the dose range of 33-66 mM during the exponential growth stage. The excision-repair-deficient strain had fewer SSB than the control group. Similar studies [124] indicated that radl_3 radiosensitive strain was more sensitive to FA than the wild-type. D N A - p r o t e i n crosslinks (DPC) were induced by FA and repair was accomplished after incubation. The induced DPC was measured by 3 ways [123], namely neutral sucrose gradient sedimentation (DPC settles at the bottom), DNA extractability (reduction in quantity of DNA), and equilibrium density gradient centrifugation (DPC causes the shift of peak to the lighter region of the gradient). (3) Drosophila melanogaster. Male Drosophila larvae were treated with FA through larva feeding

method [3]. The proposed mechanism of the mutagenicity of FA was that FA reacts with adenylic acid or adenosine, and forms N6-hydroxymethyl adenosine, an adenine ribonucleoside analogue, which acts as an antimetabolite. This antimetabolite interferes with the D N A repair of the damaged D N A in spermatocytes of the male larvae. This unique mechanism was proposed for possible application in the detection of FA induced DNA damage and repair.

(4) Mouse cell lines. Mouse leukemia cell line L1210 was utilized to compare [172,173] the degree of damage done by hexamethylmelanine (HMM), pentamethylmelanine (PMM) and FA. Both H M M and PMM are antitumor agents and believed to be FA generators when demethylized. The induced SSB and D N A crosslinks were determine by the alkaline elution method. Results indicated that FA did not induce SSB and DNA crosslinks but did promote DPC formation. Furthermore, the FA activity was not enhanced by $9 activation, and the H M M and PMM activities were not caused by FA released from the demethylization of the agents as expected. Mouse strain C3H [68] was utilized for the study of the FA effect on the D N A damage and the injury on the renal tubules. A 2% FA solution was injected into the renal cortex of the animal and observation was made on the decrease of RNA level in the renal tubules by the acridine orange staining method. RNA synthesis resumed 6 - 8 weeks after the treatment. The hypothesis was that the damaged DNA lacks the normal transcription ability, thus the RNA level was reduced. A comparative study of the crosslinkage ability of FA and potassium dichromate [50] was made. Positive results were obtained with 50 mM of FA in 2 h treatment. Both SSB and DPC were observed. (5) Rat tracheal epithelium. Doolittle [40] carried out in vivo unscheduled D N A synthesis (UDS) studies with Fisher 344 rat tracheal epithelial cell cultures. FA (0.47, 2.0, 5.9, 14.8 ppm) was utilized to fumigate the animals for 1, 3 or 5 days. The FA of this inhalation treatment failed to reach the trachea. In vitro study by administering liquid FA to the culture medium of rat tracheal epithelium

44

at 100 #M showed cytotoxicity, but no mutation or DNA damage was observed.

(6) Chinese hamster ovary (CLIO) cells.

The

CHO repair-defective line UV4, UV5 and EM9 and the wild-type AA were utilized to study the rate of DNA damage. The end point of this experiment was the growth rate of the cell culture after the treatment with same dose (5.6 #g/kg). The growth rate of the control cell line was about 2 times as high as the repair-defective lines [83].

(7) Monkey kidney cell. Experiments [142] were designed to determine the survival rate of the CV-1 kidney cells of the monkey and the ribosomal transcription rate of the cell culture after treatments with 1-16 mM of FA for 15 min. This was followed by labeling RNA with tritiated uridine. The labeled RNA extracts from the cultured kidney cell was electrophoretically separated to determine the transcription terminating lesions in the DNA template which depressed the RNA synthesis. At the 2 mM FA, 15 min treatment, DNA replication in the treated cells fell to zero. Synthesis resumed after 24 h of incubation. The UDS was not detectable in this study. (8) Human cells. Human cell cultures or cell types involves in the DNA damage/repair studies were mostly the bronchial epithelium and fibroblast cells. A series of studies carried out by Grafstr0m and his co-workers utilized FA in the dose range of 100-400 #M. The major end points for determining the FA effect was SSB, colony-forming efficiency (CFE) and DPC [63,65,66]. Comparative studies on the promutagen O6-methylguanine (OMG) [64,67], N-methyl-Nnitrosourea (MNU) and FA were made. Results indicated that FA was more mutagenic than the other agents (3 × more than MNU) and the high mutagenicity of FA was attributed to its high ability in the inhibition of repair. Comparison of the cytotoxicity between FA and X-rays (800 R) [65] and FA and v-rays (137Cs) [67] were made. There was a synergistic effect when the ionizing radiation, X-rays or ,/-rays were applied simultaneously with FA. Besides the tracheal-bronchial tissue of humans, skin fibroblasts [48,49,67], and skin tissue from xeroderma pigmentosum patients

(excision/repair-deficient) were utilized in this series of comparative studies. General response of these tissues to FA was similar to human bronchial tissue except that xeroderma pigmentosum skin tissue yielded relatively low rate of SSB [48,66]. A general mechanism of the crosslinkage of macromolecules was proposed by Harris et al. [69] through their studies of DNA damage and repair induced by N-nitrosamine. The metabolism of Nnitrosamine in human epithelium cells could generate the carbonium ions and aldehyde. Both of these metabolites may serve as alkylating agents to promote the crosslinks of DNA, DPC and SSB. These metabolites may act in concert in producing the mutagenic, clastogenic and carcinogenic effects of N-nitrosamine. In addition to the major end points used for data collection, Krokan [102] utilized the rate of inhibition of the enzymatic activities and Saladino [176] utilized the cell growth rate. The review of Weston [204] noted the used of the incorporation rate of the precursors of DNA, RNA and proteins as the measure of the effects of the FA effect on DNA in the cultured cells. Snyder and his co-workers carried out a series of studies on FA-induced DNA damage and repair, using the "nick translation" assay [186-188] in human diploid fibroblasts. The assay involved the use of tritiated nucleotides in the damage/repair synthesis and measurement of the frequencies of SSB. the end point of this assay is the radioactivity of the incorporated radioisotope-labeled nucleosides which was measured by the liquid-scintillation counting technique. Treatment with 1 /~M-100 ttM of FA (30 min) gave positive responses [186]. When the cell cultures were irradiated with 10 kRad of X-rays followed by 200 #M of FA [188], the SSB frequencies measured by the nick translation assay showed no difference in the DNA repair rate. Snyder et al., claimed that FA is ineffective in the inhibition of the repair synthesis which was contradictory to the findings of Grafstr0m's group. Coppey [30] utilized the cell survival and herpes virus production as the end points to measure the excision/repair abilities of the FA altered DNA in the skin fibroblasts of normal human subjects and that of the xeroderma pigmentosum patients. The excision/repair deficient (XP12BE) fibroblasts were about 5 times as sensitive to FA as the

45 control from normal human subjects. A similar pattern of responses were also exhibited in the herpes virus production rates in the treated and control host cells. The UV-irradiated herpes virus had lower production rate in the excision/repairdeficient host cells than in the control host cells. They claimed that there is a similarity in the DNA-repair mechanism in the UV-induced and FA-induced damage. Martin [131] utilized the HeLa cells in the study of the UDS in the FAtreated neoplastic and normal human embryo cells. The dose range of FA was between 1 × 10 -8 and 1 × 10 - 6 M with positive results based on the scintillation counting of the extracted DNA. The alkaline hydrolysate of dialyzed FA-treated RNA in which 39% of the purine nucleotides were modified to methylene his derivative ( R - C H E - R ' ) . This agent was utilized to test the cytotoxic effect in human cell culture. Cytotoxicity of these modified RNA agents was greater to human tumor cells than to the normal human embryo cells. This differential toxicity of the FA-modified RNA could be useful in the antitumor therapy [44].

III. Clastogenicity - - Chromosome damage and sister-chromatid exchange Formaldehyde was the first agent in the scientific history which induced chromosome damage in onion root tips. New interests have been raised on its clastogenic effects on human and mammalian chromosomes in vitro as well as the effects on in vivo exposure in the human population in the last decade. This renewed effort was initiated by the claim made by the Review Panel of the Interagency Regulatory Liaison Group and the Consumer Product Safety Commission which states " I t is the conclusion of the panel that it is prudent to regard formaldehyde as posing a carcinogenic risk of humans" [27]. Literature collected in this field of studies are categorized into: (1) Higher Plants, (2) Insects, (3) Amphibians, (4) Mouse, (5) Rat, (6) Hamster fibroblasts, (7) C H O and (8) Humans.

(1) Higher plants.

Vicia faba and Allium cepa

were two of the most frequently utilized plants for clastogen studies in the earlier years [11]. A current survey of literature which was published in the last decade included Tradescantia, Vicia and

Allium [116,118,121]. The Tradescantia micronucleus assay which utilizes the meiotic chromosome in the pollen mother cells as the target is a highly sensitive bioassay for gaseous [117,119, 122] and liquid [118] agents in very low concentrations. Results of tests on diesel engine exhaust fumes which is rich in FA content [118] showed positive responses at 1.5 ppm in a dynamic flow chamber for 6-h treatment, while the treatment with liquid form yielded negative results [122] but showed the signs of high toxicity to the stems. This agrees with the finding of mammalian cells culture and in vivo animal studies that FA attacks the first size of its encounter [40,75,162,180] such as the inhalation experiments in rats. The mutagenic and clastogenic actions often were reduced before it reached the deeper targets.

(2) Insects. Although Drosophila melanogaster was utilized extensively before 1977 as reported in the last review [11], there was only one publication in the recent years dealing with clastogenicity. (a) Drosophila melanogaster: No study was carried out with adult flies in this category. There were deletions observed in the salivary gland chromosomes of the larvae which were fed with FA containing medium [152]. ( b ) Chortophaga viridifasciata: The grasshopper neuroblast chromosome aberration assay is a well established test for clastogens [41]. This system has the extremely low background aberration rate, and a high sensitivity to clastogens. FA at around 30-300 ppm concentration induced 2 - 4 acentric fragments in 100 cells which were undergoing mitotic division. (3) Amphibians. The newt and frog are the best materials for monitoring pollutants in water. The larval stage of these amphibians would be the most ideal organism for teratogenicity testing because of the involvement of metamorphosis in the development to their adult life. Pleurodeles waltlii Michah: In vivo treatment of the larval stage of the newt was carried out with FA along with X-rays and other mutagens to observe the frequencies of micronuclei in the peripheral blood. The blood samples were obtained by heart puncture and the blood smear slides were

46 prepared for scoring of micronuclei. Negative resuits were obtained from the 5 ppm treated larvae but the treated animal exhibited the sign of toxicity [185]. It was probably the result of overdose of FA.

(4) Mouse. Mouse bone marrow micronucleus and sister-chromatid exchange (SCE) are both well established bioassays for clastogenicity studies. In vivo treatment by oral administration of 100 m g / k g showed the increase of MCN frequency in the polychromatic erythrocyte (PCE) from the bone marrow [156], while the lower concentrations gave negative results [139,140]. The MCN frequencies were also observed from the PCE of the spleen and the response was negative with the dosages between 6.25-25 m g / k g of FA. The resuits of SCE and MCN studies [7,22] were negative in the cases of utilizing glycerol formal as the treating agent. Kalmykova [93] found chromosome aberrations, such as translocation, ring, and chains in the meiotic spermatocytes in mice by oral administration of FA (0.3 ml FA/1) mixed with milk for 2 months. The calculated dose was around 17 m g / k g of the body weight. (5) Rats. Fisher 344 rats [97,98] were subjected to in vivo treatment with 0.5, 6 and 12 ppm of FA through inhalation. The lymphocyte samples were cultured and slides prepared for SCE and chromosome aberrations (CA). No positive result was obtained from either of these two end points.

aberrations but excluding gaps) were utilized for data collection. The FA concentration administered in the culture medium ranged from 0.1 through 0.5 g g / m l . Positive responses were obtained in both SCE and CA with total agreement between these two laboratories. This indicated the reliability of these bioassays. Obe [150] and his co-workers obtained similar positive results in C H O cultures treated with slightly higher concentration of FA, and an attempt was made to compare the effect of FA in human lymphocyte culture. Positive results on FA-induced chromosome aberrations in CHO cells were also reported by Brusick [22]. Natarajan [139] intended to make the comparison between the in vitro study of CHO-cultured cells treated with 0.006, 0.012 and 0.024 g l / m l of FA, and in vivo study with the mouse which received the treatment by i.p. injection of 6.25, 12.5 and 25.0 m g / k g of FA liquid. Both SCE and CA were utilized as the end points. A significant increase of SCE frequency was noted in the C H O cultured cells. The C H O in vitro study was carried out concurrently with the mouse (OBA strain) in vivo treatment experiment with 3 end points, namely bone marrow MCN, bone marrow CA, and spleen MCN frequencies. All in vivo tests gave negative results. No comparison between the in vitro and in vivo results could be made. Blood samples were collected 16 and 40 h after the second injection. It is highly possible that the author missed the high peak for MCN frequency in PCE if there was a slight mitotic delay.

(6) Chinese hamster lung fibroblasts (CHL). Ishidate [87-90] in his mammalian screening tests for 500 environmental agents found that FA is a weak clastogen with a T R value (a relative expression of chromosome aberration rate) of 733, while mitomycin-C had a TR value of 470000 and sodium chloride had T R value of 36. Positive response was obtained at the dosage of 7.5 g g / m l in the culture medium.

(7) Chinese hamster ovary (CHO) cells. CHOW-B1 cell line grown with rat-liver $9 activation was utilized to compare the results of two different laboratories [52], the Litton Bionetics Inc. (LBI) and Columbia University (CU). Two end points, namely SCE and CA (including chromatid

(8) Human skin fibroblasts. Out of 5 independent studies using human cell cultures, one [111] utilized skin fibroblasts, while the rest of the investigations utilized lymphocyte cultures. 4 out of 5 utilized SCE as the end points for data collection and their dosages of FA fell in the range of 0.05-300 g g / m l . One study utilized CA as the end points using the dosage of FA in the range of 60-240 g g / m l . Positive responses were obtained in the skin fibroblast [111] experiment. Dose-related increase of chromosome aberration rates were induced by 0.5-1.0 mM of FA in human lymphocytes [180], and $9 activation in the culture medium reduced the clastogenicity. The FA dosage at 2 g M had the same clastogenicity as 100 R

47 X-rays according to Levy's studies [111]. Among the SCE studies [101], FA-treated cells exhibited positive responses in 3 independent laboratories [54,101,111]. 4 independent in vivo studies of human exposures to FA were published [12,45,196,208] during the last 5 years. The FA effects through inhalation were determined by the chromosome damage or SCE in the cultured lymphocytes sampied from the formaldehyde and paper factory workers, and students of pathology and anatomy laboratories. 15 formaldehyde processing plant workers [45] were exposed to an average of 5 ppm of FA for 28 years. No significant increase of CA or Cd aberration rate was found in the exposed subjects over the control group. 6 pathology laboratory workers [196] and 5 control subjects were compared for CA, and SCE frequencies. No significant difference was found between those exposed to 1-7 ppm of FA (2-4 h/day, 2-3 days per week) for 4-11 years and the control frequency. 20 men who worked in a paper factory [12] were exposed to FA fumes at around 1 ppm (1.2 m g / m 3) for 2-30 years. The CA (dicentrics, rings) in the cultured lymphocytes were higher in the people exposed than those control subjects, and more in the longer exposure duration. Frequencies of SCE was not elevated in the exposed individuals. In an anatomy class which had around 1.2 ppm FA from the embalming solution, Yager [208] found that 8 students had slightly significant increase of SCE after 10 weeks of class than the SCE frequency observed in the cultured lymphocyte before the anatomy class.

I V. Cell transformation and carcinogenicity Most of the studies on cell transformation, tumorogenicity and carcinogenicity of FA and related aldehydes have been done in cell cultures of rodents (mouse, rats and hamsters). No publication which directly deals with normal human cell cultures was found. The study of carcinogenesis started in the late 1970s and most of the work was done in two major laboratories namely CIIT and NYU. (A) Mouse fibroblast cell line C3H/T10 1/2 was utilized for cell-transformation studies. The FA treatment (0.5-2.5 : ~ / ~ g / m l ) was usually coupled with a tumor promotor such as 12-O-

tetradecanoyl-phorbol-13-acetate (TPA) (0.1 #g/ml) [19,20]. The effective dose of FA followed by TPA was around 1.0/~g/ml (around 1 ppm in the medium) [164]. (B) Frazelle [51] treated C3H/1OT 1/2 cell culture, with N-methyl-N'-nitro-N-nitrosoguanine and FA. Transformed foci were increased by dosages (0.5-1.0/zg/ml of FA) and claimed that FA was a weak promotor for tumors. It was toxic at the dosages greater than 1.0/Lg/ml. ( C) Rat in oioo tests. Earlier inhalation (1980) studies with Fisher 344 rats and B6C3F1 mice by the CIIT group [96,191,192] at the dosages of 2, 6, 15 ppm of fumes [6 h/day, 5 day/week) for two years showed 36/240 rats and 2/240 mice with squamous carcinomas in nasal cavities at the 15 ppm concentration. Rhinitis, epithelial dysplasia and squamous metaplasia were observed in all dose levels. Tumors induced were large osteolytic neoplasms over the nasal bone. The results of inhalation studies with rats by the NYU in the early 1980s showed an even greater rate of squamous carcinomas in the nasal cavities of rats (25/100 animals), and some with benign papillomas (2/100). The average life span of the cancerous animals was around 549 days in contrast to 814 days in control rats. The synergistic effect of FA and HCI which forms bis(chloromethyl)ether (BSME) was observed in their experiments [27]. Fumigation experiments with Fisher 344 rats carried out by Rusch [175] induced squamous metaplasia and the weight loss of liver and body weight. In a follow up study of the CIIT group, Uchida [200] found no carcinoma in the nasal cavity under the similar treatment as those used by the CIIT group. Only rhinitis, squamous metaplasia and hyperplasia of nasal epithelium were noted. ( D) Rat kidney cell line HRRT was subjected to FA (10 -6 M, 3 h) and TPA a n d / o r phorbol12,13-didecanoate (PDD) tumor promotor to determine the transforming frequency [43]. Cell transformation was induced at the 1 mM concentration of FA treatment. ( E) Syrian hamster embryo ( S H E ) cells. Hatch [71,72] treated SHE with FA (2.2 ~g/ml) in liquid forms in the culture medium or gaseous forms by fumigation (3 ~1/I). Adenovirus SA7 was applied to the culture in conjunction with FA or indepen-

48 dently to determine the transforming abilities. When the adenovirus treatment was coupled with FA, the transforming ability was enhanced in the fumigation (6.0 txl/l) experiment when the sequence of virus treatment preceeded the FA fumes. Some enhancement was noted in the liquid treatment also [71,72]. Negative results were obtained when the Syrian hamsters were treated with gaseous FA (2.95 ppm) [175]. (F) Baby hamster kidney cell culture (BHK21/c1-13) was treated with FA (20 ~g/ml) and hexamethylene tetramine (HMTA) (1000 #g/ml). FA treatment exhibited high toxicity at the dosages greater than 20 ~g/ml, and a significant increase of transformed colonies in the dose range of 0.8 ~ g / m l through 20/~g/ml. HMTA is known to be a FA generator through hydrolysis (HMTA---, ammonia + FA). A significant increase of transformed colonies was noted in the dosage of 10-1000/~g/ml [159]. ( G) Cynomolgus monkeys [175] were fumigated with 2.95 ppm of FA. The only effects found were hoarseness and squamous metaplasia. N o carcinoma in the nasal cavity was observed.

V. General toxicology and teratogenic effects (A ) General toxicology Lung epithelial cells and CHO cells grown on collagen were treated with FA fumes (10-50/~g/l) for inducing morphological changes of the cultured cells [209]. Results showed shrinkage and toxicity. The effects of FA in vitro or in vivo treatment of spermatozoa of bulls and rams, and of mice and humans were studied independently [155,202]. When spermatozoa of bull and ram were preincubated in FA (0.015%) containing medium, there was an increase of glucosephosphate isomerase (GPI) but decrease of lactic dehydrogenase (LHD) in the fluid surrounding the cells [154]. The sperm morphology of the B6C3F1 strain of mice were studied after the in vivo exposure of FA (100 m g / k g / d a y ) for 5 days. No abnormal sperms were detected. In a separate study, Osinowo [155] found that 0.005% FA in vitro treatment immobilized the ram spermatozoa, and 66.7% of the immobilized sperm recovered after removal of the FA treatment. He proposed that immobilization mechanism might be related

to the membrane attachment or meta0olic inhibition. A series of studies was carried out by Sentein [181,182] and his coworkers on the disturbance of spindle fiber by FA in Triturus helvecticus Raz. and Pleurodeles waltlii Michah. Multipolar formation and segmentative mitosis were observed in the developing eggs. The proposed mechanism was based upon the FA-mediated crosslinking of protein, DPC and DNA, and the inhibition of the spindle fiber tubule formation. Electron micrographic demonstration [8,9] in this series of studies revealed that chromosomes were condensed and the spindle fiber tubules were shortened by the FA treatment. Kamata [94,95], reported that lipid peroxidase and non-protein sulfhydryl and lipid contents in the respiratory tract and lung of the Fisher 344 rats treated with FA (14.5-105.3 ppm) fumes in vivo were disturbed. There was a decrease of triglyceride and increase of phospholipids. In the review of physiological effects of FA, Gamble [53] established the 50% respiratory rate (RDs0) as the indicator of the FA effect on the respiratory tract. The physiological effects of urea-formaldehyde-foam-insulation (UFFI) was known to have the binding ability with macromolecules and promote the binding of cells [137]. The application of UFFI to houses and crops to prevent freezing should be re-evaluated.

(B) Teratogenic effects Up to date, no evidence was obtained in the teratogenic effect of FA in humans. Neither has the teratogenic effect been demonstrated in any animal species which were treated through inhalation or oral ingestion. Staples [187] and Gofmekler [58] in their inhalation experiments with rats (1 m g / m 3, 24 h/day, 10-15 days) noted the decrease in weight of fetal lung, and liver, and increase in weight of kidney and thymus gland. No histological, external malformation, and macroscopic structural changes were observed. There was increase of fetal weight but decrease of adrenal gland when rats were treated with 0.25 ml of 6% FA injections during the gestation period. Similar experiments [167] were conducted with mice (1.5 ml of 2% FA injection) during 5 h and ll.5-day gestation period, one out of 88 litters developed

49

cleft palate, and more of these abnormalities were observed when 0.3 ml of 4% FA was injected. Oral gavage treatment of CD-1 mice with 185 m g / k g / day FA showed lethal effects, but no teratogenic effects. No embryotoxic effects were seen at lower dosages [127]. Similarly, another study [82] added 125 and 375 ppm FA to the diet of beagle dogs. No adverse effects were noted in the dams or their offspring. Woodbury [206] reported the special case of 2 families each with a child born with birth defects. The mothers stayed in the U F F I insulated homes most of the time. Most teratogenic studies were done with insufficient sample size and lack of statistical reliability.

V1. Genotoxicity of related aldehydes Acetaldehyde (AA) is one of the close relatives of FA. It exists naturally in the human blood stream in trace quantities without harmful effects. Excessive concentrations could be elevated through ethanol consumption. Its genotoxicity was well established especially to the hematopoietic progenitor cell in the bone marrow. Current review of the literature is collectively categorized under the types of bioassays used. (A) Point mutation induced by AA (0.1%) in E. coli (WP2, uvrA t r p - ) was about 4.5 times higher than that of the control [84]. When bacterial culture was treated with AA plus 20-40 krad of v-rays, an additive effect was exhibited. ( B ) Sex linked recessive lethal mutation induced by AA rate was tested with standard Basc (Muller-5) assay. Positive results were obtained by the injection of AA but not by feeding [207]. (C) Crosslinks and SSB were induced by AA at the dosages of 20-49 /~M in human leukocytes [74,107] and in human bronchial epithelial cells [176]. The SSB, CPC and DNA crosslinks were observed. In human lymphocytes, Ristow [171] found that AA promotes the renealing of the damaged DNA in calf-thymus cells, and claimed that the renealing process was assisted by the crosslinking process induced by AA. ( D) Sister-chromatid exchange. A number of SCE tests were conducted to detect AA effects on chromosomes of lymphocytes, and on the bone marrow of mice [146,147,1491 hamster [99], C H O [36] and humans [18,74,146,148,149,171]. Positive responses were obtained in all these studies. Obe

[145] claimed that APt is mutagenic, not ethanol, based upon his comparative studies of AA and ethanol effect in humans. Acetaldehyde also induced the elevated frequencies of SCE in Allium root tip cell [31]. ( E ) Chromosome damage. Chromosome aberration and micronucleus bioassays were applied to the Allium [31], Vicia [170], mouse (C57B1) [120] and hamster [143] in vivo tests; on rat fibroblasts, and human lymphocytes [143] in vitro test [16]. Increased rates of chromosome aberrations and micronuclei were obtained.

(F)

Cell transformation and carcinogenesis.

Transformation studies were carried out with rat ( C 3 H / 1 0 T 1 / 2 ) cell culture under the CIIT group. Effective doses of AA were in the range of 10-25 /~g/kg. Positive results were obtained when applied in conjunction with the tumor promotor 12-O-tetradecanoylphorbol-13-acetate [1,2]. (G) Cell proliferation. A A may affect the hematopoietic progenitor cell [133], liver epithelial cell and fetal tissue cell proliferation [42] which was usually manifested by the decline of DNA synthesis or cell count. Studies on glutaraldehyde and other aldehydes such as glyceraldehyde, and malondialdehyde were carried out in bacteria, newt, rats, and cultured cells of Chinese hamsters. Salmonella tests on glutaraldehyde [73,174] and DL glyceraldehyde [179] were positive, and $9 activation again reduced the mutagenic effect of the agents as in the case of FA. Ates and Sentein [9] found that glutaraldehyde, at 1.25 mM concentration inhibited the formation of the kinetochore tubules in the mitotic chromosomes of Triturus helveticus Raz. Chromosome aberration, micronuclei and instances of aneuploidy were induced by malonaldehyde (0.1-1 mM) in rat skin fibroblasts with a positive dose-related increase. Glutaraldehyde (0.1-0.5 /xg/ml) treated C H O cells yielded positive responses in SCE tests and positive responses in polyploidy in human leukemia cell (HL60) [183].

VII. Epidemiological studies and risk assessment Since the inhalation of FA studies in rats indicated that the nasal cancer rate was increased significantly, epidemiologicai studies of cancer rate of human respiratory system was carded out in sample populations which were occupationally in-

50 volved with FA fumes. Results of two independent surveys in chemical workers [129,205] showed no significant increase of cancer rate in the respiratory system. However, the prostate cancer rate showed significant increase [205]. Although there is no clear cut link between the FA exposure and the cancer rate, epidemiological data of earlier 80s show a 0.6% increase (mainly lung cancer) in the exposed group of industrial cohorts [85]. Similarly, two surveys on the mortality of the enbalmers who were exposed to FA fumes for a number of years, showed no ill effect on the pulmonary function [110] or increased death rate due to cancer of the respiratory system including nasal cancer [201]. Nevertheless, the death rate of the arteriosclerotic heart diseases and kidney cancer were slightly elevated. In some cases, the skin cancer incidences were elevated in the workers of 35 years in the FA-contaminated atmosphere. The major epidemiological studies were reviewed by the Panel of the Consensus Workshop in the early 1980s. Most of the studies were carried out in the adult male workers in the industrial sample population. No case of nasal carcinoma has been reported. Case control studies made in Finland and Denmark showed no association between the nasal carcinoma and FA exposure. Lung cancer and mortality rate were elevated in the professional sector of the population, but not in the factory workers. Smoking might be the contributing factor in the professional sector [168]. Statistics of the workers of British Industrial Plastics showed slight increase of mortality but not solely attributed to FA exposures. No effect on morbidity, pulmonary function and vitality was noted. No quantitative assessment was made in all the epidemiological studies so far. Epidemiological studies of the British Industry Plastic workers by Infante [85] used the Standard Mortality Ratio (SMR) to express the effect of FA on lung cancer and bronchitis. The lung cancer mortality rate was in direct proportion with the FA exposure. The studies made in the common population [198], showed the seasonal variation of common symptoms of FA effects, and new symptoms were added such as skin irritation, wheezing and difficulty in breathing in the winter. Meaningful assessment can only be made with tangible results from animal studies and reliable

epidemiological data. Currently, there are sufficient evidences for the nasal carcinoma induced by the inhaled FA in rats based upon the studies of C I I T [192] and N Y U . Since rats are obligatory nose breathers whereas humans are not, it would be hard to establish the criteria to extrapolate the animal data to humans. Furthermore, the dose response of FA over carcinogenicity is non-linear and the cytotoxicity and cell production in the region of the nasal passage was dependent on the concentration of FA in the dose rate but not in the total dose derived from p p m × rain. Therefore the carcinogenicity of FA to humans can not be easily estimated [190]. Assessment was made with animal models and it was predicted that an exposure of 2 p p m would cause metaplasia in rat nasal passages and exposure of 6-15 p p m would cause hyperplasia and squamous cell carcinoma [261. N o extrapolation of the risk to humans was made [55]. The level of FA for human sensory detection is around 1 p p m [26]. Latarjet [108] established a radiation equivalent for FA doses. For lethal effect, 1 m M x 1 min of FA is equal to 6.6 rad. For chromosome aberrations, 1 m M × 1 rain is equals to 3.7 rad, and for the induction of aneuploidy, 1 m M × 1 rain is equal to 5 rad. There is no standard population threshold established so far, since some people are sensitive to very low level of exposure [86]. Currently, the standard occupational exposure limit is 3 p p m in the United States which is higher than the limit of other nations. The U.S. Occupational Safety and Health Administration estimated that the exposure of FA at the 3 p p m level for 45 years could have an excess cancer risk of 0.6%. Conclusions and discussion In the last decade, one of the major advancements in the field of FA studies is in the area of the biochemistry of its metabolism, and its reactions with macromolecules of genetic importance [8,63,69,1321. A m o n g the biochemical reactions, crosslinkage of D N A , D N A with protein and nucleoproteins are the essential mechanisms of clastogenicity and carcinogenicity as well as the subtle changes of the genetic material, point mutations.

51 Results of 30 some individual studies on point mutation in prokaryotes yielded 75% positive responses. Some of the negative results were due to the antimutagenic effect of $9 microsomes which detoxified the FA instead of providing the activation as intended. Under these circumstances, FA should be qualified as a mutagen in prokaryotes. Both yeast [25] and Neurospora [21] tests proved that FA is a potent mutagen, so did the T K mutation induction in both mouse [22] and human lymphocyte [60] cultures. At least 70 some point mutations of metabolic deficiency in humans are known. Special emphasis on the studies of these mutations and epidemiological survey on the FAinduced metabolic deficiency mutations should be made. Concerning the FA effects on DNA d a m a g e / repair and inhibition of synthesis, D N A damage was induced in all 8 different biological systems and crosslinkage of the DNAs, DNA with protein, and nucleoproteins were demonstrated in most of the mammalian cells, including extensive data obtained from studies of human cells. These findings from biological studies reconfirm the validity of the molecular crosslinks exhibited in the biochemical studies. It is understandable that FA possess such ability to link DNA and proteins because in natural living systems, FA is responsible for the biosynthesis and perhaps even abiotic synthesis of the basic subunits of these macromolecules [59]. By the same token, its omnipresence on earth as the results of industrial application should not be considered as unusual, since its presence in the universe and in the interstellar space was a natural state of creation. The problem with the FA in the modern world is that the FA fumes are now present around the living and working places at the above normal concentration. The clastogenicity of FA was manifested by chromosome and chromatid aberrations, micronuclei and SCEs. Out of 8 different in vitro and in vivo systems tested, all of the in vitro tests that administered FA directly into the culture media were positive while the in vivo tests were mostly negative with the exception of Tradecantia-micronucleus tests. This discrepancy, again, seems to be due to its action over the sites of immediate encounter. The exceptional case in Tradescantiamicronucleus test was due to the fact that the

gaseous FA was able to diffuse into the pollen mother cells readily without going through many layers of tissue or circulating systems as in living animals to transport the FA to the target cells. Results of cell transformation and carcinogenicity studies were obtained from mammalian cell cultures and live mammals respectively. Mice and rats were utilized extensively in both CIIT and N Y U inhalation experiments, and hamsters and monkeys were used in each of the independent experiments. All the transformation studies with mice, rats and hamsters cells yielded positive responses. Inhalation experiment demonstrated increased rate of squamous carcinoma in the nasal cavity of mice and rats. This is the most crucial evidence for FA to be classified as a carcinogen. Although no similar nasal cancer was observed in humans who received comparable dose of FA through occupational exposures so far, such risk should not be totally ruled out. The difference between human and rodents, as far as the route of exposure is concerned, is that the rodents are obligatory nose breathers while humans are not. Based upon the current literature survey, there is no clear cut evidence that FA has the teratogenic potential in mice, rats and beagle dogs. There is no epidemiologlcal data showing the teratogenic effect in human population so far, although there are various toxicological effects exhibited in human cell cultures. The inability of FA to induce teratological changes in embryos and fetuses is because that the target sites were well protected from the direct contact with FA. There has been a number of epidemiological studies underway since the publication of the Report of Federal Panel on FA in 1982 [169]. Emphases were on FA effects on mortality, morbidity, and the common complaints in the FA-affected population. Among the studies on the genotoxicity of related aldehydes, acetaldehyde is the only one that has the sufficient data base to be qualified as a mutagen/clastogen at the present time. Reports of the Federal Panel on FA [169] and the Consensus Workshop on FA [1681 provided a number of crucial topics for future studies. In addition to those suggestions, the following recommendations are made for the research emphases in the future.

52 ( 1 ) T h e a p p l i c a t i o n o f sensitive s h o r t - t e r m bioassays to d o the on-site m o n i t o r i n g o f low level F A is needed. This s h o u l d b e d o n e c o n c u r r e n t l y with the c h e m i c a l d e t e c t i o n a n d m e a s u r e m e n t s . N o m a t t e r h o w low a n d how a c c u r a t e l y the F A levels are d e t e c t e d b y c h e m i c a l means, it w o u l d be meaningless if the biological effects of the F A u n d e r a given e n v i r o n m e n t a l c o n d i t i o n is n o t known, especially its synergistic effects c o u p l e d with o t h e r p o l l u t a n t s a n d / o r the e n v i r o n m e n t a l factors. A p p l i c a t i o n o f b i o a s s a y p r i o r to the c h e m ical m e a s u r e m e n t s c a n help to set the p r i o r i t y schedules for e n v i r o n m e n t a l m a n a g e m e n t a n d to save some of the u n n e c e s s a r y efforts in the r o u t i n e c h e m i c a l analysis. ( 2 ) In light o f the p o t e n t i a l of F A in the i n d u c t i o n o f m u t a t i o n s in p r o k a r y o t e s which directly o r i n d i r e c t l y affect h u m a n health, a n d the s u b t l e m u t a t i o n s c o u l d b e i n d u c e d b y F A in hum a n s that w o u l d i m p a i r the i m m u n e abilities a n d n o r m a l m e t a b o l i s m , studies o f the direct links b e t w e e n c a n c e r a n d F A e x p o s u r e seems to be of s e c o n d a r y i m p o r t a n c e . F o r the far reaching p r o b lems of g e n e r a t i o n s to come, it w o u l d be m o r e m e a n i n g f u l to c a r r y on the in vitro research o f the m e t a b o l i c deficiency m u t a t i o n s in h u m a n cell cultures, a n d the e p i d e m i o l o g i c a l surveys on the imm u n e deficiency m u t a t i o n s a n d new allergic s y m p t o m s in h u m a n p o p u l a t i o n in reference to F A exposure. T h e c u r r e n t review focuses m o r e on the genetic, cytogenetic, D N A d a m a g e a n d c a r c i n o g e n i c effects of F A to all levels o f living beings i n c l u d i n g humans. Special a t t e n t i o n s h o u l d be p a i d to its subtle yet insidious actions on the vital molecules o f life. T h e o m n i p r e s e n c e of this u n i q u e single c a r b o n c o m p o u n d at the a b o v e n o r m a l c o n c e n t r a tion in o u r e n v i r o n m e n t m a y have the u n f o r e s e e a ble i m p a c t s on the ecology o f the p l a n e t earth.

Acknowledgements T h e a u t h o r s wish to express their d e e p e s t a p p r e c i a t i o n to Mr. R o b e r t S. S t a f f o r d of the Environmental Mutagen, Carcinogen and Teratogen Information Program, Oak Ridge National L a b o r a t o r y , for his assistance in searching for the references a n d the s u p p l y of a large p o r t i o n of the reprints for this review. O u r d e e p e s t g r a t i t u d e is

e x t e n d e d to the c o n t r i b u t o r s w h o r e s p o n s e d p r o m p t l y to o u r r e q u e s t for reprints, a n d the staff o f the I n t e r - L i b r a r y L o a n d e p a r t m e n t of the L i b r a r y of W e s t e r n Illinois U n i v e r s i t y for their c o o p e r a t i o n a n d assistance.

References 1 Abernethy, D.J., J.H. Frazelle and CJ. Boreiko (1982) Effects of ethanol, acetaldehyde and acetic acid in the C3H/10T1/2 C1 8 cell transformation system, Environ. Mutagen., 4, 331. 2 Abernethy, D.J., J.H. Frazeile and C.J. Boreiko (1983) Relative cytotoxic and transforming potential of respiratory irritants in the C3H/10T1/2 cell transformation system, Environ. Mutagen., 5, 419. 3 Alderson, T. (1985) Formaldehyde-induced mutagenesis: a novel mechanism for its action, Mutation Res., 154, 101-110. 4 Anonymous (1984) Formaldehyde-emitting products, Science News, 125, 106. 5 Anonymous (1981) Health hazards of formaldehyde, Lancet, 1,926-927. 6 Ashby, J., and P. Lefevre (1983) Genetic toxicology studies with formaldehyde and closely related chemicals including hexamethylphosphoramide (HMPA), in: J.E. Gibson (Ed.) Formaldehyde Toxicity, Hemisphere Publ. Corp., New York, pp. 85-97. 7 Ashby, J., and F. Ratpan (1986) Inactivity of glycerol formal in a mouse micronucleus assay: relationship to its teratogenicity, Environ. Mutagen., 8, 873-877. 8 Ates, Y., and P. Sentein (1976) Action de la formamide sur l'appareil achromatique et les chromosomes dans des oeufs de pleurodele en segmentation, Arch. Biol. (Bruxelles). 87, 367-383. 9 Ates, Y., and P. Sentein (1978) A comparison between the actions of colchicine, chloral hydrate, glutaraldehyde and phenylurethane on the ultrastructure of segmentation mitoses in a newt, Biol. Cell., 33, 129-136. 10 Attwood, M.M., and J.R. Quayle (1983) Formaldehyde as a central intermediary metabolite of methylotrophic metabolism, New Directions, 315-323. 11 Auerbach, C., M. Moutschen and J. Moutschen (1977) Genetic and cytogenetical effects of formaldehyde and related compounds, Mutation Res., 39, 317-362. 12 Bauchinger, M., and E. Schmid (1985) Cytogenetic effects in lymphocytes of formaldehyde workers of a paper factory, Mutation Res., 158, 195-199. 13 Bedford, P., and B.W. Fox (1981) The role of formaldehyde in methylene dimethanesulfonate-induced DNA cross-links and its relevance to cytotoxicity, Chem. Biol. Interact., 38, 119-126. 14 Bender, J.R., L.S. Mullin, G.J. Graepel and W.E. Wilson (1983) Eye irritation response of humans to formaldehyde, Am. Ind. Hyg. Assoc. J. 44, 463-465. 15 Benyajati, C., A.R. Place and W. Sofer (1983) Formaldehyde mutagenesis in Drosophila: molecular analysis of ADH-negative mutants, Mutation Res., 111, 1-7.

53 16 Bird, R.P., H.H. Draper and P.K. Basrur (1982) Effect of malonaldehyde and acetaldehyde on cultured mammalian cells: production of micronuclei and chromosomal aberrations, Mutation Res., 101,237-246. 17 Blackwell, M., H. Kang, A. Thomas and P. Infante (1981) Formaldehyde: evidence of carcinogenicity, Am. Ind. Hyg. Assoc. J., 42, 34-46. 18 Boehlke, J.U., S. Singh and H.W. Goedde (1983) Cytogenetic effects of acetaldehyde in lymphocytes of Germans and Japanese: SCE, clastogenic activity and cell cycle delay, Human Genet., 63, 285-289. 19 Boreiko, C.J., D.B. Couch and J.A. Swenberg (1983) Mutagenic and carcinogenic effects of formaldehyde, in: Tice, Costa and Schaich (Eds.), Environmental Science Research, Genotoxic Effects of Airborne Agents, Plenum, New York, pp. 353-367. 20 Boreiko, C.J., and D.L. Ragan (1983) Formaldehyde effects in the C 3 H / 1 0 T 1 / 2 cell transformation assay, in: J.E. Gibson (Ed.), Formaldehyde Toxicology, Hemisphere Puhl. Corp., New York, pp. 63-71. 21 Brockman, H.E., C.Y. Hung and F.J. de Serres (1981) Potent mutagenlcity of formaldehyde in a nucleotide excision repair-deficient heterokaryon of Neurospora crassa, Environ. Mutagen., 3, 379-380. 22 Brusick, D.J. (1983) Genetic and transforming activity of formaldehyde, in: J.E. Gibson (Ed.), Formaldehyde Toxicity, Hemisphere Publ. Corp., New York, pp. 72-84. 23 Casanova-Schmitz, M., and H.d'A. Heck (1983) Effects of formaldehyde exposure on the extractability of DNA from proteins in the rat nasal mucosa, Toxicol. Appl. Pharmacol., 70, 121-132. 24 Chanet, R., and R.C. Von Borstel (1979) Genetic effects of formaldehyde in yeast. 3. Nuclear and cytoplasmic mutagenic effects, Mutation Res., 62, 239-253. 25 Chaw, Y.M., L.E. Crane, P. Lange and R. Shapiro (1980) Isolation and identification of cross-links from formaldehyde-treated nucleic acids, Biochemistry, 19, 5525-5531. 26 Clary, J.J. (1983) Risk assessment for exposure to formaldehyde, in: J.E. Gibson (Ed.), Formaldehyde Toxicity, Hemisphere Publ. Corp., New York, pp. 284-294. 27 Coene, R.F. (1981) Formaldehyde: evidence of carcinogenicity, Vet. Human Toxicol., 23, 282-285. 28 Conner, T.H., M.D. Battle, J.C. Theiss, T.S. Matney and J.B. Ward Jr. (1983) Mutagenicity of formalin in the Ames assay, Mutation Res., 119, 145-149. 29 Conner, T.H., J.C. Theiss, H.A. Hanna, D.K. Monteith and T.S. Mamey (1985) Genotoxicity of organic chemicals frequently found in the air of mobile homes, Toxicol. Lett., 25, 33-40. 30 Coppey, J., and D. Nocentini (1979) Survival and herpes virus production of normal and xeroderma pigmentosum fibroblasts after treatment with formaldehyde, Mutation Res., 62, 355-361. 31 Cortes, F., S. Mateos and P. Escalza (1986) Cytotoxic and genotoxic effects of ethanol and acetaldehyde in rootmeristem cells of Allium capa, Mutation Res., 171, 139-143. 32 Couch, D.B., P.F. Allen and H.C. Eales (1982) The mutagenicity of formaldehyde to Salmonella typhimurium, Environ. Mutagen., 4, 336-337.

33 De Flora, S. (1981) study of 106 organic and inorganic compounds in the Salmonella/microsome test,. Carcinogenesis, 2, 283-298. 34 De Flora, S., A. Camoirano, P. Zanacchi and C. Bennicelli (1984) Mutagenicity testing with TA97 and TA102 of 30 DNA-damaging compounds, negative with other Salmonella strains, Mutation Res., 143, 159-165. 35 De Flora, S., P. Zanacchi, A. Camoirano, C. Bennicelli and G.S. Badolati (1984) Genotoxic activity and potency of 135 compounds in the Ames reversion test and in a bacterial DNA-repair test, Mutation Res., 133, 161-198. 36 De Raat, W.K., P.B. Davis and G.L. Bakker (1983) Induction of sister-chromatid exchanges by alcohol and alcoholic beverages after metabolic activation by rat-liver homogenate, Mutation Res., 124, 85-90. 37 De Serres, F.J. (1983) The use of Neurospora in the evaluation of the mutagenic activity of environmental chemicals, Environ. Mutagen., 5, 341-351. 38 De Serres, F.J. (1986) Genetic characterization of the mutagenic activity of environmental chemicals at specific loci in two-component heterokaryons of Neurospora crassa, in: C. Ramel, B. Lambert, J. Magnusson (Eds.), Progress in Clinical and Biological Research Series, 209A, Genetic Toxicology of Environmental Chemicals, Part A, Basic Principles and Mechanisms of Action, Liss, New York, pp. 200-218. 39 Donovan, S.M., D.F. Krahn, J.A. Stewart and A.M. Sarrif (1983) Mutagenic activities of formaldehyde (HCHO) and hexamethylphosphoramide (HMPA) in reverse and forward Salmonella typhimurium mutation assay, Environ. Mutagen., 5, 476. 40 Doolittle, D.J., and B.E. Butterworth (1984) Assessment of chemically-induced DNA repair in rat tracheal epithelial cells, Carcinogenesis, 5, 773-779. 41 Dowd, M.A., M.E. Gaulden, B.L. Proctor and G.B. Seibert (1986) Formaldehyde induced acentric chromosome fragments and chromosome stickiness in Chortophaga neuroblasts, Environ. Mutagen., 8, 401-411. 42 Dreosti, I.E., F.J. Ballard, G.B. Belling, I.R. Record, S.J. Manuel and S.S. Hetzel (1981) The effect of ethanol and acetaldehyde on DNA synthesis in growing cells and on fetal development in the rat, Alcoholism, Clin. Exptl. Res., 5, 357-362. 43 Eker, P., and T. Sanner (1986) Initiation of in vitro cell transformation by formaldehyde and acetaldehyde as measured by attachment-independent survival of cells in aggregates, Eur. J. Cancer Clin. Oncol., 22, 671-676. 44 Feldman, M.Y., H. Balabanova, U. Bachrach and M. Pyshnov (1977) Effect of hydrolized formaldehyde-treated DNA on neoplastic and normal human cells, Cancer Res., 37, 501-506. 45 Fleig, I., N. Petri, W.G. Stocker and A.M. Thiess (1982) Cytogenetic analysis of blood lymphocytes of workers exposed to formaldehyde in formaldehyde manufacturing and processing, J. Occup. Med., 24, 1009-1012. 46 Fontignie-Houbrechts, N. (1981) Genetic effects of formaldehyde in the mouse, Mutation Res., 88, 109-114. 47 Fontignie-Houbrechts, N., J. Moutschen, M. MoutschenDahraen, N. DeGrave and H. GIoor (1982) Genetic effects in the mouse of formaldehyde in combination with

54

48

49

50

51

52

53

54

55

56

57

58

59

60

61

62

63

64

adenosine and hydrogen peroxide, Mutation Res., 104, 371-376. Fornace, Jr., A.J. (1982) Detection of DNA single-strand breaks produced during the repair of damage by DNA-protein cross-linking agents, Cancer Res., 42, 145-149. Fornace Jr., A.J., J.F. Leclmer R.C. GrafstriSm and C.C. Harris (1982) DNA repair in human bronchial epithelial cells, Carcinogenesis, 3, 1373-1377. Fornace Jr., A.J., D.S. Seres, J.F. Lechner and C.C. Hams (1981) DNA-protein cross-linking by chromium salts, Chem. Biol. Interact., 36, 345-354. Frazelle, J.H., D.J. Abernethy and C.J. Boreiko (1983) Weak promotion of C H 3 H / 1 0 T 1 / 2 cell transformation by repeated treatments with formaldehyde, Cancer Res., 43, 3236-3239. Galloway, S.M., A.S. Bloom, M. Resnick, B.H. Margolin, F. Nakamura, P. Archer and E. Zeiger (1985) Development of a standard protocol for in vitro cytogenetic testing with Chinese hamster ovary cells: comparison of resuits for 22 compounds in two laboratories, Environ. Mutagen., 7, 1-51. Gamble, J. (1983) Effects of formaldehyde on the respiratory system, in: J.E. Gibson (Ed.), Formaldehyde Toxicity, Hemisphere Publ. Corp., New York, pp. 175-195. Garry, V.F., R.A. Kreiger and J.K. Wiencke (1981) The mutagenic and cytogenic effects of formaldehyde in cultured human lymphocytes, Environ. Mutagen., 3, 341. Gaylor, D.W. (1983) Mathematical approaches to risk assessment: Squamous cell carcinoma in rats exposed to formaldehyde vapor, in: J.E. Gibson (Ed.), Formaldehyde Toxicity, Hemisphere Publ. Corp., New York, pp. 279-283. Glass, L.R., T.H. Connor, J.C. Theiss, C.E. Dallas and T.S. Matbey (1986) Genotoxic evaluation of the offgassing products of particle board, Toxicol. Lett., 75-83. Gocke, E., M.-T. King, K. Eckhardt and D. Wild (1981) Mutagenicity of cosmetic ingredients licensed by the European communities, Mutation Res., 90, 91-109. Gofmekler, V.A. (1968) Effect on embryonic development of benzene and formaldehyde in inhalation experiments, Hyg. Sanit., 33, 327-332. Goldberg, L. (1983) Forward, in: J.E. Gibson (Ed.), Formaldehyde Toxicology, Hemisphere Publ. Corp., New York, pp. 17-19. Goldberg, L. (1983) Risk assessment with formaldehyde: Introductory remarks, in: J.E. Gibson (Ed.), Formaldehyde Toxicology, Hemisphere Publ. Corp., New York, pp. 275-277. Goldmacher, V.S., P. Temcharoen and W.G. Thilly (1982) Mutagenic effects of formaldehyde in bacterial and human cells, in: Clary, Gibson and Waritz (Eds.), Formaldehyde: Toxicology, Epidemiology and Mechanisms, Marcel Dekker, New York. pp. 173-191. Goldmacber, V.S., and W.G. Thilly (1983) Formaldehyde is mutagenic for cultured human cells, Mutation Res., 116, 417-422. Grafstrt~m, R.C., R.D. Curren, L.L., Yang and C.C. Harris (1985) Genotoxicity of formaldehyde in cultured human bronchial fibroblasts. Science, 228, 89-91. Grafstr~Sm, R.C., R.D. Curren, L.L. Yang and C.C. Hams

65

66

67

68

69

70

71

72

73

74

75

76

77 78

(1986) Aldehyde-induced inhibition of DNA repair and potentiation of N-nitroso compound-induced mutagenesis in cultured human cells, in: C. Ramel, B. Lambert and J. Magnusson (Eds.), Progress in Clinical and Biological Research Series, 209A Genetic Toxicology of Environmental Chemicals, Part A: Basic principles and Mechanisms of Action, Liss, New York, pp. 255-264. Grafstr/Sm, R.C., A.J. Fornace Jr., H. Autrup, J.F. Lechner and C.C. Hams (1983) Formaldehyde damage to DNA and inhibition of DNA repair in human bronchial cells, Science, 220, 216-218. GrafstrtSm, R.C., A. Fornace Jr. and C.C. Hams (1984) Repair of DNA damage caused by formaldehyde in human cells, Cancer Res., 44, 4323-4327. Grafstr/Sm, R.C., A.E. Pegg, C.C. Hams, K. Sundqvist and H. Ka'okan (1985) Ot-Methylguanine-DNA methyltransferase activity and aldehyde induced inhibition of Ot-methylguanine repair in human lung cells, in: Myrnes and Krokan (Eds.), Repair of DNA Lesions Introduced by N-Nitroso Compounds, Norwegian University Press, Oslo, pp. 154-175. Greenberg, S.R. (1982) Nucleic acid relationships in formalin-injured renal tubules, Proc. Inst. Med. (Chicago) 35, 83-84. Hams, C.C., R.C. GrafstrOm, J.F. Lechner and H. Autrup (1982) Metabolism of N-nitrosamines and repair of DNA damage in cultured human tissues and cells, in: Magee (Ed.), Nitrosamines and Human Cancer, Banbury Report 12, Cold Spring Harbor Laboratory, New York, pp. 121-139. Hams, J.C., B.H. Rumack and F.D. Aldrich (1981) Toxicology of urea formaldehyde and polyurethane foam insulation, J. Am. Med. Assoc., 245, 243-246. Hatch, G.G. and T.M. Anderson (1985) increased virus transformation by formaldehyde of hamster embryo cells pretreated with benzo{a]pyrene, in: Waters, Sandhu, Lewtas, Claxton, Strauss and Nesnow (Eds.), Short-term Bioassays in the Analysis of Complex Environmental Mixtures, Vol. IV, Plenum, New York, pp. 125-138. Hatch, G.G., P.M. Conklin, C.C. Christensen, B.C. Casto and S. Nesnow (1983) Synergism in the transformation of hamster embryo cells treated with formaldehyde and adenovirus, Environ Mutagen., 5, 49-57. Haworth, S., T. Lawlor, K. Mortelmans, W. Speck and E. Zeiger (1983) Salmonella mutagenicity test results for 250 chemicals, Environ. Mutagen., 5, 3-142. He, S.M., and B. Lambert (1985) Induction of persistence of SCE-inducing damage in human lymphocytes exposed to vinyl acetate and acetaldehyde in vitro, Mutation Res., 158, 201-208. Heck, H. d'A., and M. Casanova-sehrnitz (1984) Biochemical toxicology of formaldehyde, Rev. Biochem. Toxicol. 6, 155-189. Heck, H. d'A., T.Y. Chin and M.C. Schmitz (1983) Distribution of 14C-formaldehyde in rats after inhalation exposure, in: J.E. Gibson (Ed.), Formaldehyde Toxicity, Hemisphere Publ., New York, pp. 26-37. Hemminki, K. (1981) reaction of formaldehyde with guanosine, Toxicol. Lett., 9, 161-164. Hemminki, K. (1982) Urinary sulfur containing metabo-

55

79 80 81

82

83

84

85 86

87

88

89 90

91

92

93 94

95

iites after administration of ethanol acetaldehyde and formaldehyde to rats, Toxicol. Lett., 11, 1-6. Hemminki, K. (1982) Dimetbylnitrosamine adducts excreted in rat urine, Chem.-Biol. Interact., 39, 139-148. Hemminki, K. (1984) Urinary excretion products of formaldehyde in the rat, Chem.-Biol. Interact., 48, 243-248. Hileman, B. (1982) Formaldehyde: How did EPA develop its formaldehyde policy, Environ. Sci. Technol., 16, 543A-547A. Hoy, C.A., E.P. Salazar and L.H. Thompson (1984) Rapid detection of DNA-damaging agents using repair-deficient CHO cells, Mutation Res., 130, 321-332. Humi, H., and H. Ohden (1973) Reproduction study with formaldehyde and hexamethylenetetramine in beagle dogs, Food Cosmet. Toxicol., 11,459-462. Igali, S., and L. Gazso (1980) Mutagenic effect of alcohol and acetaldehyde on Escherichia coli, Mutation Res., 74, 209-210. Infante, P.F., and M.A. Schneiderman (1986) Formaldehyde, lung cancer and bronchitis, Lancet, Feb. 22, 436-437. lnfante, P.F., A.G. Ulsamer, D. Groth, K.C. Chu and J. Ward (1981) Health hazards of formaldehyde, Lancet, 2, 980-982. ishidate Jr., M. (1981) Application of chromosomal aberration tests in vitro to the primary screening for chemicals with carcinogenic a n d / o r genetic hazards, in: Short-term Tests for Carcinogenesis: Quo Vadis? Proceedings of a symposium held in Montpellier on Feb. 4-5, Excerpta Medica, Amsterdam, pp. 58-79. Ishidate Jr., M. (1982) Mammalian cell systems as a screening tool for the detection of possible mutagens and carcinogens in the environment, EISEI Kagaku, J. Hyg. Chem., 28, 291-304. Ishidate, M. (1987) Chromosomal Aberration Test in vitro, Monograph Realize, Inc. Ishidate Jr., M., T. Sofuni and K. Yoshikawa (1981) Chromosomal aberration tests in vitro as a primary screening tool for environmental mutagens a n d / o r carcinogens, Gann, Monograph Cancer Res., 27, 95-108. Jeffcoat, A.R., F. Chasalow, D.B. Feldman and H. Mart (1983) Disposition of t4C-formaldehyde after topical exposure to rats, guinea pigs, and monkeys, In: Gibson (Ed.) Formaldehyde Toxicity, Hemisphere Publ. Corp., New York, pp. 38-49. Jones, A.R., and P.M. Mashford (1983) The fate of Nm e t h y I - N ' - ( h y d r o x y m e t h y l ) t h i o u r e a in the rat, Xenobiotica, 13, 73-79. Kalmykova, T.P. (1979) Cytogenetic effect of formalin on somatic and germ cells of animals, Veterinariya, 11, 67-69. Kamata, E., O. Uchida, S. Suzuki, Y. Ikeda, Y. Ogawa, T. Kaneko and M. Tobe (1984) Toxicological effects of some inhaled chemicals on the respiratory tract and the lungs of rats (first report), J. Toxicol. Sci., 9, 303. Kamata, E., O. Uchida, S. Suzuki, Y. Ikeda, Y. Ogawa, T. Kaneko and M. Tobe (1985) Toxicological effects of some inhaled chemicals on the respiratory tract and the lungs of the rats, (2nd report), J. Toxicol. Sci., 10, 258.

96 Kerns. W.D., D.J. Donofrio, K.L. Pavkov (1983) The chronic effects of formaldehyde inhalation in rats and mice: A preliminary report, in: J.E. Gibson (Ed.), Formaldehyde Toxicity, Hemisphere Publ. Corp., New York, pp. 111-131. 97 Kligerman, A.D., M.C. Phelps and G.L. Erexson (1984) Cytogenetic analysis of lymphocytes from rats following formaldehyde inhalation, Toxicol. Lett., 21,241-246. 98 Kligerman, A.D., J.L. Wilmer and G.L. Erexson (1983) Analyses of cytogenetic damage in rat lymphocytes after inhalation of toxicants, Environ. Mutagen., 5, 400. 99 Korte, A., G. Obe, I. lngwersen and G. Rueckert (1981) Influence of chronic ethanol uptake and acute acetaldehyde treatment on the chromosome of bone marrow cells and peripheral lymphocytes of Chinese hamsters, Mutation Res., 88, 389-395. 100 Kosako, M., and H. Nishioka (1982) New forward mutation assay using low-concentration streptomycin resistance mutation in E. coli strains with plasmid PKM101, Sci. Eng. Rev. Doshisha University, 22, 239-249. 101 Kreiger, R.A., and V.F. Garry (1983) Formaldehyde-induced cytotoxicity and sister-chromatid exchanges in human lymphocyte cultures, Mutation Res., 120, 51-55. 102 Krokan, H., R.C. GrafstrSm, K. Sundqvist, H. Esterbauer and C.C. Harris (1985) Cytotoxicity, thiol depletion and inhibition of O6-methylguanine-DNA methyltransferase by various aldehydes in cultured human broncl~ial fibroblasts, Carcinogenesis, 6, 1755-1759. 103 Kubinski, H., G.E. Gutzke and Z.O. Kubinski (1981) DNA-cell binding (DCB) assay for suspected carcinogens and mutagens, Mutation Res., 89, 95-136. 104 Kubinski, H., and Z.O. Kubinski (1986) Practical and theoretical aspects of the DCB assay for carcinogenic and other genotoxic agents, Toxicity Assessment: Int. Q., 1, 387-406. 105 Kubinski, H., Z.O. Kubinski and J. Shahidi (1985) Use of formalin-treated cells in the DNA-cell binding assay for genotoxic and carcinogenetic chemicals, in: Li (Ed.), New Approaches in Toxicity Testing and Their Application in Human Risk Assessment, Raven, New York, pp. 93-97. 106 L'Abbe, K.A., and J.R. Hoey (1984) Review of the health effects of urea-formaldehyde foam insulation, Environ. Res., 35, 246-263. 107 Lambert, B.O., Y.U. Chen, S.-M. He and M. Sten (1985) DNA cross-links in human leukocytes treated with acetate and acetaldehyde in vitro, Mutation Res., 146, 301-303. 108 Latarjet, R. (1982) Quantitative comparison of genotoxic (mutagenic and carcinogenic) risks and the choice of energy sources, Bull. Acad. Natl. Med. (Paris) 166, 181-201. 109 Levin, D.E., M. HoUstein, M.F. Christman, E.A. Schwiers and B.N. Ames (1982) A new Salmonella tester strain (TA102) with A-T base pairs at the site of mutation detects oxidative mutagens, Proc. Natl. Acad. Sci. (U.S.A.) 79, 7445-7449. 110 Levine, R.J., R.D. Dal Corso, P.B. Blunden and M.C. Battigelli (1983) The effects of occupational exposure on

56 the respiratory health of West Virginia morticians, in: J.E. Gibson (Ed.), Formaldehyde Toxicity, Hemisphere Publ. Corp., New York, pp. 212-226. 111 Levy, S., S. Nocentini and C. BiUardon (1983) Induction of cytogenetic effects in human fibroblast cultures after exposure to formaldehyde or X-rays, Mutation Res., 119, 309-317. 112 Lewis, B.B., and S.B. Chestner (1981) Formaldehyde in dentistry: a review of mutagenic and carcinogenic potential, J. Am. Dental Assoc., 103, 429-434. 113 Liu, K.S., S.B. Hayward, G. Kulasingam, B.H. Chang and K. Sexton (1986) Estimation of formaldehyde exposure for mobile home residents, 79th Ann. Meet. Air Pollut. Control Assoc., Minneapolis, MN, June 22-27, Voi. 8668.1, pp. 1-15. 114 Liu, K.S., K. Sexton, S.B. Hayward, M. Petreas, L. Webber and B.H. Chang (1986) Determinants of formaldehyde concentrations inside mobile homes, 79th Ann. Meet. Air Pollut. Control Assoc., Minneapolis, MN, June 22-27, Vol. 86-7.7, pp. 1-16. 115 Loomis, T.A. (1979) Formaldehyde toxicity, Arch. Pathol. Lab. Med., 103, 321-324. 116 Ma, T.H., and M.M. Harris (1985) In situ monitoring of environmental mutagens, in: Saxena (Ed.), Hazard Assessment of chemicals, Current Developments, Academic Press, New York, pp. 77-106. 117 Ma, T.H., and M.M. Harris (1987) Tradescantia-micronucleus (Trad-MCN) bioassay - - A promising indoor air pollution monitoring system,. 4th International Conf. on Indoor Air Quality and Climate, West Berlin, Aug. 17-21, pp. 243-247. 118 Ma, T.H., M.M. Harris, V.A. Anderson, I. Ahmed, K. Mohammad and J.L. Bare (1984) Tradescantia-micronucleus (Trad-MCN) tests on 140 health-related agents, Mutation Res., 138, 157-167. 119 Ma, T.H., M.M. Harris and G. Lin (1985) Genotoxicity studies of formaldehyde using Tradescantia-micronucleus test, Environ. Mutagen., 7, 42. 120 Ma. T.H., M.M. Harris, Z. Xu and T.L. Miller (1985) Genotoxic effect of acetaldehyde detected by Mouse-peripheral erythtrocyte micronucleus test, Proc. 4th International Conference of Environmental Mutagens, Stockholm, Sweden, p. 55. 121 Ma, T.H., Z. Xu, L.G. Cabrera, R.M.E. Valtiera and G.G. Arreola (1987) The efficacy of root tip cell micronucleus assays for water pollutants, Environ. Mutagen., 9, 65. 122 Ma, T.H., Z. Xu, M.M. Harris and G. Lin (1986) Dose response of formaldehyde fume-induced chromosome damage in Tradescantia pollen mother cells, Environ. Mutagen., 8, 49, 123 Magana-Schwencke, N., and B. Ekert (1978) Biochemical analysis of damage in yeast by formaldehyde, 2. Induction of cross-links between DNA and protein, Mutation Res., 51, 11-19. 124 Magana-schwencke, N., B. Ekert and E. Moustacchi (1978) Biochemical analysis of damage induced in yeast by formaldehyde. 1. Induction of single-strand breaks in DNA and their repair, Mutation Res., 50, 181-193.

125 Magana-Schwencke, N., E. Moustacchi (1980) Biochemical analysis of damage induced in yeast by formaldehyde, 3. Repair of induced cross-links between DNA and proteins in the wild-type and in excision-deficient strains, Mutation Res., 70, 29-35. 126 Marison, I.W., and M.M. Attwood (1982) A possible alternative mechanism for the oxidation of formaldehyde to formate, J. Gen. Microbiol., 128, 1441-1445. 127 Marks, T.A., W.C. Worthy and R.E. Staples (1980) Influence of formaldehyde and Sonacide (potentiated acid glutaraldehyde) on embryo and fetal development in mice, Teratology, 22, 51-58. 128 Marnett, L.J., H.K. Hurd, M.C. Hollstein, D.E. Levin, H. Esterbauer and B.N. Ames (1985) Naturally occurring carbonyl compounds are mutagens in Salmonella tester strain TA104, Mutation Res., 148, 25-34. 129 Marsh, G.M. (1983) Proportionate mortality among workers, in: J.E. Gibson (Ed.), Formaldehyde Toxicity, Hemisphere Publ. Corp., New York, pp. 237-254. 130 Marshall, E. (1987) EPA indicts formaldehyde, 7 years later, Science, 236, 381. 131 Martin, C.N., A.C. McDermid and R.C. Garner (1978) Testing of known carcinogens and noncarcinogens for their ability to induce unscheduled DNA synthesis in HeLa cells, Cancer Res., 38, 2621-2627. 132 Mashford, P.M., and A.R. Jones (1982) Formaldehyde metabolism by the rat: a re-appraisal, Xenobiotica, 12, 119-124. 133 Meagher, R.C., F. Sieber and J.L. Spivak (1982) Suppression of hematopoietic-progenitor-cell proliferation by ethanol and acetaldehyde, N. Engl. J. Med., 307, 845-849. 134 Miller, A.J., J.W. Pensabene, R.C. Doerr and W. Fiddler (1985) Apparent mutagenicity of N-nitrosothiazolidine caused by a trace contaminant, Mutation Res., 157, 129-134. 135 Mitsevich, E.V., Yu.A. Semin, A.M. Poverennyi and Yu.G. Suchkov (1979) Effect of formaldehyde and its aminomethylol derivatives on strains of Escherichia coli with various defects of DNA repair systems, Bull. Exptl. Biol. Med. (U.S.S.R.), 87, 489-491. 136 Moerman, D.G., and D.L. Baillie (1981) Formaldehyde mutagenesis in the nematode Caenorhabditis elegans, Mutation Res., 80, 273-279. 137 Morin, N.C., and H. Kubinski (1978) Potential toxicity of materials and for home insulation; Ecotoxicol. Environ. Safety, 2, 133-141. 138 Nasrat, G.E., A.H. Abdel-Rahman, H.A. Nafei and K.A. Ahmed (1977) The effect of temperature on the mutagenic action of formaldehyde and maleic hydrazide on Drosophila melanogaster, Bull. Faculty Agri., Univ. Cairo, 28, 719-734. 139 Natarajan, A.T., F. Darroudi, C.J.M. Bussman and A.C. Van Kesteren-Van Leeuwen (1983) Evaluation of the mutagenicity of formaldehyde in mammalian cytogenetic assays in vivo and vitro, Mutation Res., 122, 355-360. 140 Natarajan, A.T., and G. Obe (1986) How do in vivo mammalian assays compare to in vitro assays in their ability to detect mutagens? Mutation Res., 167, 189-201.

57 141 Newell, G. (1983) Overview of formaldehyde, in: J.E. Gibson (Ed.), Formaldehyde Toxicity, Hemisphere Publ. Corp., New York, pp. 3-12. 142 Nocentini, S., G. Moreno and J. Coppey (1980) Survival, DNA synthesis and ribosomal RNA transcription in monkey kidney cells treated by formaldehyde, Mutation Res., 70, 231-240. 143 Norppa, H., F. Tursi, P. Pfaeffle, J. Maeki-Paakkanen and H. Jaerventans (1985) Chromosome damage induced by vinyl acetate through in vitro formation of acetaldehyde in human lymphocytes and chinese hamster ovary cells, Cancer Res., 45, 4816-4821. 144 Norsted, S.W., C.A. Kozinetz and J.F. Annegers (1985) Formaldehyde complaint investigations in mobile homes by the Texas Department of Health, Environ. Res., 37, 93-100. 145 Obe, G. (1981) Acetaldehyde not ethanol is mutagenic, Progress in Mutation Res., 2, 19-23. 146 Obe, G. (1981) Mutagenicity and carcinogenicity of alcobol, Mutation Res., 85, 221-222. 147 Obe, G., and B. Beek (1979) Mutagenic activity of aldehydes, Drug and Alcohol Dependence, 4, 91-94. 148 Obe, G., R. Jonas and S. Schmidt (1986) Metabolism of ethanol in vitro produces a compound which induces sister chromatid exchanges in human peripheral lymphocytes in vitro: acetaldehyde not ethanol is mutagenic, Mutation Res., 174, 47-51. 149 Obe, G., A.T. Natarajan, M. Meyers and A. Den Hertog (1979) Induction of chromosomal aberrations in peripheral lymphocytes of human blood in vitro, and of SCEs in bone marrow cells of mice in vivo by ethanol and its metabolite acetaldehyde, Mutation Res., 68, 291-294. 150 Obe, G., and H. Ristow (1977) Acetaldehyde, but not ethanol, induces sister chromatid exchanges in chinese hamster cells in vitro, Mutation Res., 56, 211-213. 151 Oda, Y., S. Nakamura, I. Oki, T. Kato and H. Shinagawa (1985) Evaluation of the new system (UMU-Test) for the detection of environmental mutagens and carcinogens, Mutation Res., 147, 219-299. 152 O'Donnell, J., H.C. Mandel, M. Krauss and W. Sofer (1977) Genetic and cytogenetic analysis of the ADH region in Drosophila melanogaster, Genetics, 86, 553-566. 153 Ohba, Y., Y. Morimitsu and A. Watarai (1979) Reaction of formaldehyde with calf-thymus nucleohistone, Eur. J. Biochem., 100, 285-293. 154 Osinowo, O.A. (1981) Studies on leakage of enzymes from washed bull and ram spermatozoa, J. Reprod. Fert., 62, 549-554. 155 Osinowo, D.A., J.O. Bale, E.O. Dyedipe and L.D. Eduvie (1982) Motility and eosin uptake of formaldehyde-treated ram spermatozoa, J. Reprod. Fert. 65, 389-394. 156 Pereira, M.A., L.W. Chang, L. McMillan, J.B. Ward and M.S. Legator (1982) Battery of short-term tests in laboratory animals to corroborate the detection of human population exposures to genotoxic chemicals, Environ. Mutagen., 4, 317. 157 Pickrell, J.A., B.V. Molder, L.C. Griffis, C.H. Hobbs and A. Bathija (1983) Formaldehyde release rate coefficients

158

159

160

161

162

163

164

165

166

167 168 169 170

171

172

173

174

from selected consumer products, Environ. Sci. Technol., 17, 753-757. Place, A.R., C. Benyajati and W. Sorer (1982) The molecular consequences of formaldehyde and ethyl methanesulfonate mutagenesis in Drosophila: analysis of mutants in the alcohol dehydrogenase gene, in: Vl71nga (Ed.), Methods in Protein Sequence Analysis, Humana Press, Clifton, N J, pp. 373-379. Plesner, B.H., and K. Hansen (1983) Formaldehyde and hexamethylenetetramine in Styles' cell transformation assay, Carcinogenesis, 4, 457-459. Polacow, I., L. Cabasso and H.J. Li (1976) Histone redistribution and conformational effect on chromatin indued by formaldehyde, Biochemistry, 15, 4559-4565. Pool, B.L., and M. Wiessler (1981) Investigations on the mutagenicity of primary and secondary alphaacetoxynitrosamines with Salmonella typhimurium activation and deactivation of structurally related compounds by S-9, Carcinogenesis, 2, 991-997. Proctor, B.L., M.E. Gaulden and M.A. Dowd (1986) Reactivity and fate of benzene and formaldehyde in culture medium with and without fetal calf serum; relevance to in vitro mutagenicity testing, Mutation Res., 160, 259-266. Protosova, I.A., Y.A. Semin, V.V. Alder and A.M. Poverennyi (1982) Influence of formaldehyde and products of its interaction with amines on process of DNA transcription in vitro, Biochemistry (U.S.S.R.), 47, 1509-1515. Ragan, D.L., and C.J. Boreiko (1981) Initiation of C 3 H / 1 0 T 1 / 2 cell transformation by formaldehyde, Cancer Left., 13, 325-331. Randerrath, K., E. Randerrath, H.P. Agarwal, R.C. Gupta, M.E. Schurdak and M.V. Reddy (1985) Postlabeling methods for carcinogen-DNA adduct analysis, Environ. Health Perspect., 62, 57-65. Randerrath, K., M.V. Reddy and R.C. Gupta (1981) 32p. labeling test for DNA damage, Proc. Natl. Acad. Sci. (U.S.A.), 78, 6126-6129. Ranstroem, S. (1956) Stress and pregnancy, Acta Pathol. Microbiol Scand. Suppl., 111,113-114. Report on the Consensus Workshop on Formaldehyde (1984) Environ. Health Perspect., 58, 323-381. Report of the Federal Panel on Formaldehyde (1982) Environ. Health Perspect., 43, 139-168. Rieger, R., and A.O. Michaelis (1960) Chromosome aberrations after action of acetaldehyde on primary roots of Vicia faba, Biol. Zbl. (Leipzig), 79, 1-5. Ristow, H., and G. Obe (1978) Acetaldehyde induces cross-links in DNA and causes sister chromatid exchanges in human cells, Mutation Res., 58, 115-119. Ross, W.E., D.R. McMillan and C.F. Ross (1981) Comparison of DNA damage by methylmelamines and formaldehyde, J. Natl. Cancer Inst., 67, 217-221. Ross, W.E., and N. Shipley (1980) Relationship between DNA damage and survival in formaldehyde-treated mouse cells, Mutation Res., 79, 277-283. Ruiz-Rubio, M., E. Alejandre-Duran and C. Pueyo (1985) Oxidative mutagens specific for A-T base pairs induce

58 forward mutations to L-arabinose resistance in Salmonella typhimurium, Mutation Res., 147, 153-163. 175 Rusch, G.M., J.J. Clary, W.E. Rinehart and H.F. BoRe (1983) A 26-week inhalation toxicity study with formaldehyde in the monkey, rat and hamster, Toxicol. Appl. Pharmacol., 68, 329-343. 176 Saladino, A.J., J.C. Willey, J.F. Lechner, R.C. GrafstriSm, M. LaVeck and C.C. Harris (1985) Effect of formaldehyde, acetaldehyde, benzoyl peroxide and hydrogen peroxide on cultured normal human bronchial epithelial cells, Cancer Res., 45, 2522-2526. 177 Salas, L.J., and H.B. Singh (1986) Measurements of formaldehyde and acetaldehyde in the urban ambient air, Atm. Environ., 20, 1301-1304. 178 Sarrif, A.M., S.M. Donovan, J.A. Stewart, D.F. Krahn and J.F. Russell Jr. (1983) Inhibition by semicarbazide of the mutagenic activities of formaldehyde (HCHO) and hexamethylphosphoramide (HMPA) in Salmonella typhimurium reverse suspension assay, Environ. Mutagen., 5, 476. 179 Sasaki, Y., and R. Eldo (1978) Mutagenicity of aldehydes in Salmonella, Mutation Res., 54, 251-252. 180 Schmid, E., W. Goggelmann and M. Bauchinger (1987) Formaldehyde-induced cytotoxic, genotoxic and mutagenic response in human lymphocytes and Salmonella typhimurium, Mutagenesis, 1, in press. 181 Sentein, P. (1976) Action of several amides on the achromatic apparatus and the chromosomes in cleaving Urodele eggs, Cytobiologie, 12, 305-320. 182 Sentein, P., and Y. Ates (1978) Cytological characteristics and classification of spindle inhibitors according to their effects on segmentation mitoses, Cellule, 72, 267-289. 183 Sentein, P., and D.L. Coq (1986) Enhanced polyploidy by glutaraldehyde in cultured HL60 leukemic cells, Leukemia Res., 10, 575-584. 184 Sexton, K., K.S. Liu and M.X. Petreas (1986) Formaldehyde concentrations inside private residences: a mail-out approach to indoor air monitoring, J. Air Pollut. Control Assoc., 36, 698-704. 185 Siboulet, R., S. Grinfeld, P. Deparis and A. Jaylet (1984) Micronuclei in red blood cells of the newt Pleurodeles waltlii Michah: induction with X-rays and chemicals, Mutation Res., 125, 275-281. 186 Snyder, R.D. (1986) Evaluation of putative inhibitors of DNA excision repair in cultured human cells by the rapid nick translation assay, Mutation Res., 173, 279-286. 187 Snyder, R.D., and D.W. Matheson (1985) Nick translation a new assay for monitoring DNA damage and repair in cultured human fibroblasts, Environ. Mutagen., 7, 267-279. 188 Snyder, R.D., and B. Van Houten (1986) Genotoxicity of formaldehyde and an evaluation of its effects on the DNA repair process in human diploid fibroblasts, Mutation Res., 165, 21-30. 189 Staples, R.E. (1983) Teratogenicity of formaldehyde, in: J.E. Gibson (Ed.), Formaldehyde Toxicity, Hemisphere Publ. Corp., New York, pp. 51-59. 190 Swenberg, J.A., C.S. Barrow, C.J. Boreiko, H.d'A Hock, -

-

R.J. Levine, K.T. Morgan and T.B. Starr (1983) Non-linear biological responses to formaldehyde and their implications for carcinogenic risk assessment, Carcinogenesis, 4, 945-952. 191 Swenberg, J.A., C.A. Gross, J. Martin and J.A. Popp (1983) Mechanisms of formaldehyde toxicity, in: Gibson (Ed.), Formaldehyde Toxicity, Hemisphere Publ. Corp., New York, pp. 132-147. 192 Swenberg, J.A., W.D. Kerns, R.I. Mitchell, E.J. Gralla and K.L. Pavkov (1980) Induction of squamous cell carcinomas of the rat nasal cavity by inhalation exposure to formaldehyde vapor, Cancer Res., 40, 3398-3402. 193 Szabad, J., I. Sots, G. Polgar and G. Hejja (1983) Testing the mutagenicity of malondialdehyde and formaldehyde by the Drosophila mosaic and the sex-linked recessive lethal tests, Mutation Res., 113, 117-133. 194 Takahashi, K., T. Morita and Y. Kawazoe (1985) Mutagenic characteristics of formaldehyde on bacterial systems, Mutation Res., 156, 153-161. 195 Temcharoen, P., and W.G. Thilly (1983) Toxic and mutagenic effects of formaldehyde in Salmonella typhimurium, Mutation Res., 119, 89-93. 196 Thomson, E.J., S. Shackleton and J.M. Harrington (1984) Chromosome aberrations and sister-chromatid exchange frequencies in pathology staff occupationally exposed to formaldehyde, Mutation Res., 141, 89-93. 197 Thun, M., and R. Altman (1983) Symptom survey of residents of homes insulated with urea-formaldehyde foam: methodologic issues, in: J.E. Gibson (Ed.), Formaldehyde Toxicity, Hemisphere Publ. Corp. New York, pp. 198-202. 198 Thun, M.J., M.F. Lakat and R. Ahman (1982) Symptom survey of residents of homes insulated with urea-formaldehyde foam, Environ. Res., 29, 320-334. 199 Tweats, D.J., M.H.L. Green and W.J. Muriel (1981) A differential killing assay for mutagens and carcinogens based on an improved repair-deficient strain of Escherichia coil, Carcinogenesis, 2, 189-194. 200 Uchida, O., E. Kamata, M. Saito, Y. Ikeda, Y. Ogawa, T. Kaneko and M. Tobe (1984) Inhalation toxicity study of formaldehyde (Nasal cavity cancer of rats), J. Toxicol. Sei., 9, 303. 201 Walrath, J., and J.F. Fraumeni, Jr. (1983) Proportionate mortality among New York embalmers, in: J.E. Gibson (Ed.), Formaldehyde Toxicity, Hemisphere Publ. Corp., New York, pp. 227-235. 202 Ward Jr., J.B., J.A. Hokanson, E.R. Smith, L.W. Chang, M.A. Pereira, E.B. Whorton Jr. and M.S. Legator (1984) Sperm count, morphology and fluorescent body frequency in autopsy service workers exposed to formaldehyde, Mutation Res., 130, 417-424. 203 Wartew, G.A. (1983) The health hazards of formaldehyde, J. Appl. Toxicol., 3, 121-126. 204 Weston, A., S.M. Plummer, R.C. Grafstr/Sm, B.F. Trump and C.C. Harris (1986) Genotoxicity of chemical and physial agents in cultured human tissues and cells, Food Chem. Toxicol., 24, 675-679. 205 Wong, O. (1983) An epidemiologic mortality study of a

59 cohort of chemical workers potentially exposed to formaldehyde, with a discussion on SMR and PMR, in: J.E. Gibson (Ed.), Formaldehyde Toxicity, Hemisphere Publ. Corp., New York, pp. 256-271. 206 Woodbury, M.A., and C. Zenz (1983) Formaldehyde in the home environment: prenatal and infant exposures, in: J.E. Gibson (Ed.), Formaldehyde Toxicity, Hemisphere Publ. Corp., New York, pp. 203-210. 207 Woodruff, R.C., J.M. Mason, R. Valencia and S. Zimmering (1985) Chemical mutagenesis testing in Drosophila, 5. Results of 53 coded compounds tested for the National Toxicology Program, Environ. Mutagen., 7, 677-702.

208 Yager, J.W., K.L. Chon, R.C. Spear, J.M. Fisher and L. Morse (1986) Sister-chromatid exchanges in lymphocytes of anatomy students exposed to formaldehyde-embalming solution, Mutation Res., 174, 135-139. 209 Zamora, P.O., J.M. Benson, T.C. Marshall, B.V. Molder, A.P. Li, A.R. Dahl, A.L. Brooks and R.O. McClellan (1983) Cytotoxicity and mutagenicity of vapor-phase pollutants in rat lung epithelial cells and Chinese hamster ovary cells grown on collagen gels, J. Toxicol. Environ. Health, 12, 27-38.