Increased frequency of chromosome aberrations in workers exposed to styrene.

and SKERFVING, S. Increased frequency of chromosome aberrations in workers exposed to styrene. Scand. j. work environ. & health 5 (1979) 333-335. Chromosomal aberrations in lymphocytes from peripheral blood were significantly more frequent in six workers from a plant manufacturing polyester resin boats (average 10.8 per 100 cells) than in six age- and sex-matched referents (5.2 per 100 cells). The contam ination of the workroom air with styrene, as measured on three occasions within three years in different areas of the plant, was 50-400 mg/m 3.

Styrene and styrene oxide, which is an intermediate metabolite of styrene, have proved mutagenic in a series of test systems (2,5,6,7,9,12). In workers exposed to styrene an increased frequency of chromosomal aberrations in lymphocytes has been reported in studies of considerably exposed groups (3,8), while no such effect has been found in a slightly exposed one (11). In these studies the degree of contamination was uncertain, as it was estimated from single determinations of styrene metabolites in the urine. This paper reports cytogenetic findings concerning workers from a plant in which the contamination of the workroom air

Production
The study was performed in a small plant manufacturing fiberglass-reinforced polyester resin boats. The polyester precursors were phthalic acid anhydride, maleic acid anhydride, propylene glycol, and styrene. The hardener used was methyl ethyl ketone peroxide, and the catalyst was a cobalt salt. Acetone was used for cleaning purposes. The production started in the late 1960s. The gel coat and laminate were sprayed, while the top coat was rolled on. Ventila-0355-3140/79/040333-3 tors, although rather primitive, were installed before 1973, and spray boxes in 1976-1977.

Measurements of contamination
The styrene levels in the air were measured on several occasions in individual samples from the breathing zones of workẽ rs. In 1971 samples were obtained in impinger flasks (ethanol and 0.1 ppm tertiary buthyl pyrocathecol). In 1973 and 1977 samples were collected in charcoal tubes and eluted by carbon disulfide. In 1973 urine sampLes, obtained at the end of a day's work, were analyzed spectrophotometrically (10) for styrene metabolites (mandelic acid and phenylglyoxylic acid) and creatinine.

Subjects
The exposed group consisted of six males, aged 21-56 (mean 33) years. They had been employed at the plant for 0.5-10 (mean 4) years. The six age-and sexmatched referents were selected from workers at a nearby paper factory where there was no contamination with chemicals. Both groups were questioned regarding smoking habits, diseases (especially virus infections), use of drugs, and exposure to ionizing radiation, heavy metals, and organic solvents. Three of the exposed subjects and three of the referents were smokers. No other agents possibly capable of damaging chromosomes were known.

Cytogenetic methods
Venous blood specimens were collected in connection with the atmospheric samples in 1977. The samples from both groups were obtained and analyzed simultaneously. Lymphocyte cultures were incubated about 20 h after the collection of the blood samples. A standard microculture technique was used. The cultivation medium was a mixture of 80 % McCoy's 5a and 20 % fetal calf serum. Phytohemagglutinin was used as the mitogen substance. The cultivation time was 72 h. All preparations were coded, mixed, and analyzed by one observer. Two hundred metaphases from each individual were analyzed. The chromosomal aberrations were recorded according to the classification system recommended by the World Health Organization In 1973 the mandelic acid concentration in the urine of nine workers ranged from 225 to 2,100 (mean 490) mg/g of creatinine and that of phenylglyoxylic acid from 82 to 560 (mean 205) mg/g of creatinine.
The average frequency of different types of aberrations was higher for the exposed group than for the referents ( fig. 1). This result applied to gaps (3. There was no obvious relation between exposure time and total number of aberrations. Although the present material is small, it clearly shows that the manufacture of polyester resin boats is associated with an increased frequency (approximatelya doubling) of chromosomal aberrations. This phenomenon is presumably due to exposure to styrene although the possibility of exposure to other chemicals may have contributed to the increase in the frequency of the aberrations. The concentrations of styrene were approximately at, or even below, the maximum limits for the substance (Sweden 170 mg/ m 3 ).
The finding of an increased frequency of chromosomal aberrations is in accord with earlier findings. The present results agree closely with those published by Fleig and Thiess (3). Compared with the findings given by Meretoja et al. (8), our frequencies of aberrations were lower (especially when compared with those of the referents), despite the probably higher level (as judged from the atmospheric levels of styrene and urinary levels of mandelic acid) and longer duration of exposure in our study.
The exposure levels measured in the present study are common in many plastic industries. Thus the finding of an increased frequency of chromosomal aberrations at these levels may have serious implications. The possibility of cancer must be borne in mind, although no epidemiologic evidence is available. There are indications of an embryotoxic effect in man (4), an effect which may have a cytogenetic background. In addition there is a risk of a genotoxic effect on germ cells that could possibly not become clinically manifest for several generations.

ACKNOWLEDGMENTS
This investigation was supported by grants from the Swedish Cancer Society.
The measurements of the contamination of the air with styrene in 1971 were made by Mr. S. Seehusen.
Received for publication: 26 June 1979