Toxicology Letters 72 (1994) 205-211

Xxicology letters

Biological monitoring of environmental exposure to polycyclic aromatic hydrocarbons;

1-hydroxypyrene in urine of people

Frans J. Jongeneelen

IndusTox Consult, PO Box 31070, Nl-6503 CB Nijmegen, The Netherlands

(Accepted 24 November 1993)

A biomarker of human exposure to chemical agents provides a valuable parameter for assessing the extent and significance of the uptake by giving a measurement that is direct and integrated over time and exposure routes. Urinary I-hydroxypyrene is currently tested as a biomarker for the assessment of low level environmental exposure of people to polycyclic aro- matic hydrocarbons (PAH). Five examples of the application of urinary I-hydroxypyrene methodology in the assessment of environmental exposure to PAH are presented: inhalation of tobacco smoke; inhalation of urban outdoor air; windsurfers sailing on polluted water; ab- sorption of contaminated food; exposure in an urban area with many heavy industries. The examples illustrate that the urinary 1-hydroxypyrene test is sufIiciently sensitive. Urinary l- hydroxypyrene is an effective biomarker for the assessment of human environmental exposure to PAH.

Key wora!r: Polycyclic aromatic hydrocarbons; I-Hydroxypyrene; Biomarker; Environmental exposure

1. lo-00

Population exposure to polycyclic aromatic hydrocarbons (PAH) from various environmental sources may be direct (e.g. food, smoking, air pollution), but may

also occur as indirect exposure, for instance by contact with polluted soil or by con- tact with polluted water.

l Cormponding author. Tel:. +31 80 528842; Fax: +31 80 540090.

0378-4274t94607.00 0 1994 Elsevier Science Ireland Ltd. All rights reserved. SSDI 0378-4274(93)03091-7

206 F. J. Jongeneelen / Toxicol. Lat. 72 (1994) 205-21 I

The assessment of environmental exposure to environmental pollutants requires the measurement of levels present in each pathway of possible contact. A biomarker of exposure provides a valuable method of assessing the extent and significance of the human contact by giving a measurement that is direct and integrated over time and exposure routes.

I-Hydroxypyrene (a major metabolite of pyrene) in urine has been suggested as a biological indicator of exposure to polycyclic aromatic hydrocarbons (PAH). Several industrial hygiene studies have shown that I-hydroxypyrene in urine is a valid and sound biomarker for occupational exposure to PAH [l-4].

Currently, urinary 1-hydroxypyrene is tested as biomarker for the assessment of low level environmental exposure of people. Five examples of assessment of en- vironmental exposure to PAH are presented:

1. PAH uptake due to smoking and/or environmental tobacco smoke. 2. Uptake of PAH due to inhalation of urban outdoor air. 3. Exposure of PAH among windsurfers sailing on polluted water. 4. Oral absorption of PAH due to contaminated food. 5. Environmental exposure to PAH in an urban area with many heavy industries.

2. PAH uptake due to active smoking and/or to environmental tobacco smoke

It is well known that tobacco smoke contains PAH. Mainstream smoke contains lo-50 ng benzo(u)pyrene/cigarette and the PAH concentration of side-stream smoke is about 4 times higher than main-stream smoke [5]. A sensitive biomarker of PAH exposure should be able to detect PAH uptake due to smoking and, if possible, due to passive smoking. Several studies among inhabitants from Europe and Asia show an increased level of urinary 1-hydroxypyrene among smokers, in most studies significantly increased compared with non-smokers (Table 1).

In an experimental study with 10 volunteers the uptake of tobacco smoke consti- tuents due to passive smoking was studied. Among other biomarkers, l-hydroxy- pyrene in urine was determined. Smokers showed significantly increased urinary concentrations of 1 -hydroxypyrene, whereas the group exposed to environmental tobacco smoke did not show significantly increased levels [6].

It seems that PAH uptake due to smoking can easily be assessed with the hydroxy- pyrene biomarker, but uptake due to environmental tobacco smoke is much more difficult to detect.

3. Uptake of PAH due to inhalation of urban outdoor air

An urban environment may contain substantial airborne concentrations of PAH, especially in wintertime, due to domestic heating with coal-burning stoves. Zhao et al. [7,8] showed that outdoor air concentrations of PAH in the winter period in a business and residential area in Beijing (China) are increased 5 to 20 times and exposure to this air pollution results in significant increased excretion of urinary 1-hydroxypyrene in the winter period (21 out of 28 residents had increased l- hydroxypyrene concentrations in wintertime and the mean of the individual differ-

F. J. Jongeneelen / Toxicol. Lat. 72 (1994) 205-211 201

Table 1 Urinary I-hydroxypyrene concentrations of smoking and non-smoking controls

Group - country I-Hydroxypyrene in urine (pmol/mol creatinine) Median (number of persons)

Non-smoking Smoking P Reference

University controls - Netherlands Industrial controls - Netherlands Urban controls - China University controls - Netherlands University controls - Turkey Industrial controls - Denmark Controls - Germany

0.26 (52) 0.28 (38) 0.07 HI 0.17 (14) 0.51 (28) 0.003 121 0.68* (74) 0.76. (84) 0.33 171 0.12 (39) 0.25 (37) 0.0001 I121 0.24* (15) 0.33* (14) 0.01** [I41 0.16* (20) 0.26* (26) 0.01** 1151 0.05 (10) 0.22 (10) - 1161

*Arithmetic mean. **P-value of ANOVA of smoking among controls and exposed workers.

ence was 0.4 pmol/mol creatinine). Also a clear relationship was found between urinary I-hydroxypyrene and ambient air concentration of benzo(a)pyrene in per- sons from different urban areas [4,7].

The data show that the urinary 1-hydroxypyrene is a promising biomarker for monitoring of PAH exposure of urban residents due to inhalation of polluted air.

4. Exposure of PAH among windsurfers sailing on polluted water

Exposure to chemical agents of people recreating at polluted river water due to dermal contact and absorption of river water has, as yet, hardly been studied. The sediment in rivers of the Rhine delta in The Netherlands is highly polluted. We studied the exposure to PAH of windsurfers recreating on the ‘Ketelmeer’. The ‘Ketelmeer’ is one of the polluted waters in the Rhine delta in The Netherlands (con- centration of 16 EPA-PAH in sediment = 4.0 mg/kg dry matter). The uptake of PAH among windsurfers was established in a study of 6 persons windsurfing for 2 con- secutive days on the ‘Ketelmeer’.

The 3 days before windsurfing were reference days, the following 2 days were ‘windsurfing days’ with at least 3 h of surfing per day for the 6 volunteers. The 6 persons were all using a wet suit on the ‘windsurfing days’. The surfers collected a urine sample daily at 18:00 h. After an interval of 13 days a fourth reference urine sample was collected. The study design included the control of variation of PAH intake by smoking, ingestion, occupation, medication and inhalation.

The excretion of urinary I-hydroxypyrene on the 2 exposure days was statistically significantly increased compared to reference days (P = 0.001, Table 2), but the increased concentrations were within the range of external referents [9]. The calcula- tion of inhalatory, oral and dermal dose rates of pyrene made plausible that the major route of uptake of PAH was dermal.

The study showed that urinary I-hydroxypyrene is a biomarker that is sufficiently sensitive for the assessment of dermal uptake of PAH among those involved in watersports.

208

Table 2

F.J. Jongeneelen/ Toxicol. Lat. 72 (1994) 205-211

I-Hydroxypyrene in urine of windsurfers on days of surfing and on reference days

I-Hydroxypyrene in urine (pmol/mol creatinine)

Mean (range)

Surfing days 0.32 (0.16-0.81) Reference days 0.11 (0.05-o. 16)

The difference is statistically significant (P < 0.091).

Number of surfers

6 6

5. Oral absorption of PAH due to contaminated food

Dietary intake of PAH may be substantial. Environmental pathway studies indi- cate that diet is a main source of PAH [lo]. An experimental study with 5 volunteers consuming low-PAH meals and high-PAH meals showed that a lOO- to 250-fold increase in a dietary benzo(u)pyrene dose was parallel to a 4- to 12-fold increase in 1 -hydroxypyrene excretion in urine [ 111.

In a second study the effect of lifestyle and personal factors on I-hydroxypyrene excretion was studied. Male volunteers (n = 76) were interviewed using question- naires. The individual base-line of urinary I-hydroxypyrene was determined as the mean of 3 samples. It was concluded that smoking and dietary PAH intake were significant determinants of urinary hydroxypyrene [ 121. The dietary exposure to PAH is substantial and urinary I-hydroxypyrene holds promise as an biomarker of dietary PAH intake.

6. Environmental exposure to PAH in an urban area with many heavy industries

Upper Silesia is an area in Poland with concentrated heavy industries such as cokeries, coal-tired power plants and domestic coal heating. The area is regarded as a highly polluted area in Europe. Exposure to PAH of residents of this area might take place due to: (i) inhalation of polluted air; (ii) dermal uptake due to contact with soil; (iii) consumption of contaminated home-grown vegetables.

Table 3 I-Hydroxypyrene in urine from boys and girls from the town of Bytom in the region of Upper Silesia (Poland)

I-Hydroxypyrene

Mean (SD.) Mean (SD.)

(mnoW @mol/mol creatinine)

Boys (n = 72) 7.55 (9.31) 0.66 (0.83) Girls (n = 76) 6.25 (4.94) 0.59 (0.47)

Mean (S.D.) concentration of creatinine for boys and girls is not different: 11.5 (3.8) and 1 I .2 (3.7) mmol/l, respectively.

F. J. Jongeneelen / Toxicol. Lat. 72 (1994) 205-21 I 209

Table 4 Analysis of variance of I-hydroxypyrene in urine from 148 children of the town of Bytom in the region of Upper Silesia (Poland)

Source I-Hydroxypyrene concentration expressed as:

nmol/l ~moVmo1 creatinine

log(nmoVl) log(~mollmol creatinine)

F-value P F-value P F-value P F-value P

Sub-area 1.75 0.03 1.75 0.04 1.54 0.08 1.68 0.05 Sex 3.69 0.06 1.24 0.27 2.60 0.11 1.41 0.23

A cross-sectional study among 148 children from the town of Bytom in Upper Silesia was performed to study total environmental exposure to PAT-I. The children were 8 or 9 years old, both boys and girls and were recruited from 24 different subareas of the town of Bytom (total area of the town is 83 km2). From each subarea 5-9 children were selected. Urine was collected and the concentration of 1-hydroxypyrene was expressed as the urine concentration and as the creatinine- corrected concentration.

After sorting the data by subarea and sex some interesting results were clear. Table 3 shows the data by sex. The mean (and S.D.) concentration of urinary l-hydroxy- pyrene in girls was lower, suggesting a lower exposure. The mean 1-hydroxypyrene concentration in urine of children from the 24 subareas ranged from 2.5 11 nmol/l (= 0.3-0.8 pmol/mol creatinine). A 2-way analysis of variance was performed for the sources’ sex and subarea. The analysis of variance was tested with uncorrected l- hydroxypyrene, with the corrected 1-hydroxypyrene and with the logarithmic value of both. The results are summarized in Table 4. The analysis confirmed the

Table 5 Urinary I-hydroxypyrene concentration of children of the town of Bytom (Poland) and non-smoking adults from The Netherlands

Bytom children (Poland)

Nijmegen University adults [l] (The Netherlands)

Number Mean age (S.D.) % nude I-Hydroxypyrene in urine (nmol/l)

Median Pz~-P,~ (interquartile range) Range

I-Hydroxypyrene in urine (nmoVmmo1 creatinine) Median Pzs-P7s (intequartile range)

Range

148 52 8.5 (1) 35 (11)

51% 100%

5.07 3.21 3.34-8.51 1.74-5.05 0.70-74.1 0.05-9.36

0.46 0.26 0.32-0.74 0.14-0.41 0.09-6.99 0.005-0.81

The ditference in I-hydroxypyrene between both groups is statistically significant (P < 0.001).

210 F. J. Jongeneelen / Toxicol. Letr. 72 (1994) MS-21 I

statistically significant effect of the source subarea, but the effect of sex was not sig- nificant. 1 -Hydroxypyrene concentrations of referents (non-smoking adults from The Netherlands [l]) were available. Table 5 shows the urine concentration of the ‘Bytom children’ and the reference group. The table shows that urinary l- hydroxypyrene of ‘Bytom children’ is significantly higher. The results show that the uptake of PAH varies significantly over the town of Bytom and that the base-line uptake of children in Bytom is increased. The study did not allow the quantification of the importance of each of the exposure routes (inhalation, dermal absorption and food contamination).

7. Discussion

The presented studies illustrate the validity of urinary 1-hydroxypyrene as a biomarker of environmental exposure to PAH. This biomarker can trace low level exposure to environmental PAH. A great advantage is that the determination of urinary 1-hydroxypyrene is an easy, rapid and a non-laborious test, which makes it very suited to large scale epidemiological studies.

A disadvantage of the 1-hydroxypyrene methodology is that 1-hydroxypyrene is a metabolite of only one PAH and solely indicates exposure to pyrene. However, the partial pyrene content in the total PAH contamination is rather constant, therefore it may be regarded as an indicator of PAH.

Risk assessment of total PAH uptake using urinary 1-hydroxypyrene as bio- marker is still beyond the border of science; a relation between urinary l-hydroxy- pyrene and long-term effects from epidemiological studies is lacking. However, an indirect dose-response relationship of urinary I-hydroxypyrene versus relative risk of lung cancer in cokeoven workers has been estimated [ 131. Since the profile of PAH in the environment may deviate from the PAH profile of the cokeoven a cor- rection factor is necessary before this dose-response relationship can be applied in the estimation of long-term risks of environmental PAH exposure.

8. Acknowledgement

The fruitful co-operation of Dr. D. Mielzynska of the Institute of Occupational Medicine in Sosnowiec, Poland is gratefully acknowledged.

9. References

1 Jongeneelen, F.J. (1987) Biological monitoring of occupational exposure to PAH. Ph.D thesis, Uni- versity of Nijmegen.

2 Jongeneelen, F.J., van Leeuwen, F.E., Oosterink, S. et al. (1990) Ambient and biological monitoring of cokeoven workers; determinants of the internal dose of PAH. Br. J. Ind. Med. 47, 454-461.

3 Buchet, J.P., Gennart, J.P., Mercado-Calderon, F., Delavignette, J.P., Cupers, L. and Lauwerys, R. (1992) Evaluation of exposure to PAH in a coke production and graphite electrode manufacturing plant. Br. J. Ind. Med. 49, 761-768.

4 Zhao, Z., Quan, W. and Tian, D. (1990) Urinary I-hydroxypyrene as an indicator of human expo- sure to ambient polycyclic aromatic hydrocarbons in a coal burning environment. Sci. Total En- viron. 92, 145-154.

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5 IARC (International Agency for Research of Cancer). (1986) Monograph on the evaluation of car- cinogenic risks to humans; tobacco smoking, Vol. 38, IARC, Lyon.

6 Scherer, G., Conze, C., Tricker, A.R. and Adlkofer, F. (1992) Uptake of tobacco constituents on exposure to environmental tobacco smoke. Clin. Invest. 70, 352-367.

7 Zhao, Z., Quan, W. and Tian, D. (1992) Experiments on the effects on the I-hydroxypyrene level in human urine as an indicator of exposure to PAH. Sci. Total Environ. 113, 197-207.

8 Zhao, Z., Quan, W. and Tian, D. (1992) The relationship between PAH in ambient air and l- hydroxypyrene in human urine. J. Environ. Sci. Health, A27, 1949-1966.

9 Weerdt, R. vd. and Jongeneelen, F.J. (1991) Opname van PAK door windsurfers. T. Sot. Gezond- heidsz 69, 466-472 (in Dutch).

10 Hattemer-Frei, H.A. and Travis, CC. (1991) Benzo(a)pyrene; environmental partitioning and human exposure. Toxicol. Ind. Health 7, 141-157.

1 I Buckley, T.J. and Lioy, P.J. (1992) An examination of the time course from human dietary exposure to PAH to urinary elimination of I-hydroxypyrene. Br. J. Ind. Med. 49, 113-124.

12 VanRooij, J.G.M., Veeger, M., Bodelier-Bade, M.M. and Jongeneelen, F.J. (1994) Smoking and dietary PAH as sources of interindividual variability of urinary I-hydroxypyrene. Int. Arch. &cup. Environ. Health (in press).

13 Jongeneelen, F.J. (1992) Biological exposure limit for occupational exposure to coal tar pitch volatiles at cokeovens. Int. Arch. Qccup. Environ. Health 63, 51 l-516.

14 Burgaz, S., Bonn, P.J.A. and Jongeneelen, F.J. (1992) Evaluation of urinary I-hydroxypyrene and thioethers in workers exposed to bitumen fumes. Int. Arch. Occup. Environ. Health 63, 397-401.

15 Sherson, D., Sigsgaard, T., Overgaard, E., Loft, S., Poulsen, H.E. and Jongeneelen, F.J. (1992) In- teraction of smoking, uptake of PAH and cytochrome-450IA2 activity among foundry workers. Br. J. Ind. Med. 49, 197-202.

16 Angerer, J., Schaller, K.H. and Hausmann, N. (1991) Determination of I-hydroxypyrene as a tool for biological monitoring of PAH-exposed persons. Presentation at the 13th International Sym- posium on PAH, Bordeaux, France.