Experimental human exposure to toluene

Summary The relationship between the individual toluene uptake and the urinary hippuric acid excretion was studied under experimental conditions. Six healthy male subjects were exposed to various concentrations in inspired air (50, 100, 125, 150, and 200 ppm) at rest or under different levels of physical effort.The hippuric acid excretion near the end of the exposure appeared under all circumstances directly proportional to the time-weighted uptake rate of toluene. The correlation between respiratory uptake rate and the rate of metabolite excretion near the end of the exposure period proved not to be systematically influenced by personal factors such as body weight, amount of body fat, urine flow rate and urinary pH. The relatively pronounced differences in background excretion of hippuric acid and, perhaps, distribution phenomena of toluene between different tissues under heavy workload conditions, can partly explain the greater variability in metabolite excretions as compared to the individual uptake rates.The correlation between the individual uptake rate of toluene and the hippuric acid excretion proved substantially better when using the end exposure excretion rate as exposure parameter as compared with the end exposure hippuric acid concentration, even after correcting the latter for urine density.Reasonable biological limit values complying to an acceptable time-weighted toluene dose were found to be 3000–3500 mg/l and 2.0–2.5 mg/min, resp. for average hippuric acid concentrations and excretion rates in spot samples during the second half of a complete work shift.     

Urinary hippuric acid concentration after occupational exposure to toluene.

The results of industrial investigations have shown a correlation between the rate of hippuric acid excretion in a single urine sample collected after daily occupational exposure and the amount of toluene absorbed. The rate of hippuric acid excretion and the average concentration of toluene vapour during exposure time were also related. The quantitative range of the test has been limited to amounts exceeding 425 mg of toluene and concentrations exceeding 69 ppm of toluene in the air because of the physiological presence of hippuric acid in urine. The rate of hippuric acid excretion in urine depends on diuresis and is constant for urinary fractions with diuresis of 30 ml/h. The physiological excretion rate was 20 mg/h with a standard deviation +/- 4.3 mg/h, maximal physiological level 33 mg/h.     

Toxicokinetics of toluene and urinary excretion of hippuric acid after human exposure to 2H8-toluene.

Nine male volunteers were exposed to 2H8-toluene (200 mg/m3 for two hours during a workload of 50 W) via inspiratory air with the aid of a breathing valve and mouthpiece. Labelled toluene was used to differentiate between hippuric acid originating from exposure to toluene and hippuric acid normally excreted in urine. The total uptake of toluene was 2.2 (standard deviation (SD) 0.2) mmol, or 50% of the amount inhaled. Four hours after the end of exposure 1.4 (SD 0.3) mmol or 65% of the total uptake had been excreted in urine as 2H-hippuric acid and 20 hours after the end of exposure the cumulative excretion of 2H-hippuric acid was 1.8 (SD 0.3) mmol, or 78% of the total uptake. By contrast the cumulative excretion of labelled plus unlabelled hippuric acid exceeded the total uptake of toluene already after four hours. The excretion rate of 2H-hippuric acid was highest, about 5 mumol/min, during exposure and the SD between the subjects was low. The background concentrations of unlabelled hippuric acid in urine were high, however, and there were large differences between subjects. These findings confirm earlier indications that for low exposure, urinary hippuric acid concentration cannot be used for biological monitoring of exposure to toluene.     

Three fatal cases of thinner-sniffing,and experimental exposure to toluene in human and animals

Summary Three fatal cases of organic solvent abuse revealed high levels of toluene in blood and alveolar air and a high level of hippuric acid, metabolite of toluene, in urine. The lethal concentration of toluene was estimated to be 2,000 ppm.Furthermore, 10 male and female volunteer students were exposed to 107 ±12 ppm toluene for 4 hours. Hippuric acid in urine increased with the exposure time and reached maximum 2 hours after initiation of toluene exposure and remained at the same level thereafter. Following cessation of exposure to toluene, hippuric acid in urine showed a rapid decrease and recovered almost to the normal level 4 hours after cessation of exposure.Urinary excretion of hippuric acid in 7 rabbits exposed to 350 ppm for 100 minutes or to 4,500 ppm toluene for 10 minutes, reached its maximum 1.5–2 hours after initiation of exposure and decreased rapidly after cessation of exposure to toluene to recover to the normal level 4 hours later.Read before the 43rd Annual Meeting of Japanese Association of Industrial Health at Tokushima on April 2, 1970, and the 18th Annual Meeting of North Kanto Medical Association at Maebashi on November 14, 1971.     

Effects of ethanol and phenobarbital treatments on the pharmacokinetics of toluene in rats.

Rats were exposed to toluene at a wide range of concentrations from 50 to 4000 ppm for six hours, and the effects of ethanol and phenobarbital (PB) treatments on the pharmacokinetics of toluene metabolism were investigated. Ethanol treatment influenced toluene metabolism mainly at low exposure concentrations. Thus ethanol accelerated the clearance of toluene from blood only when the blood concentration of toluene was not high (less than 360 microM), and ethanol increased hippuric acid (HA) excretion in urine more significantly at low (less than 250 ppm) than at high atmospheric toluene concentrations. Ethanol also expressed a similar effect on p-cresol excretion as on HA, but had little effect on o-cresol. Phenobarbital treatment promoted the urinary excretion of all of the metabolites of toluene, especially after exposure to high toluene concentration. As well as HA, benzoylglucuronide (BG) and free benzoic acid were found in urine. These are the products of the side chain metabolism of toluene. Amounts of BG could be detected when the urinary excretion of free benzoic acid exceeded 5 mumol/kg/6 h, indicating that a great deal of benzoic acid is required for the formation of BG. The Michaelis constant (Km) and the maximum rate of metabolic excretion in urine during six hours exposure (Vmax) of isozymes involved in the excretion of toluene metabolites were calculated, and correlated with the subtypes of cytochrome P-450. The significance of the result was suggested in the biological monitoring of exposure to toluene.     

Effects of ethanol and phenobarbital treatments on the pharmacokinetics of toluene in rats.

Rats were exposed to toluene at a wide range of concentrations from 50 to 4000 ppm for six hours, and the effects of ethanol and phenobarbital (PB) treatments on the pharmacokinetics of toluene metabolism were investigated. Ethanol treatment influenced toluene metabolism mainly at low exposure concentrations. Thus ethanol accelerated the clearance of toluene from blood only when the blood concentration of toluene was not high (less than 360 microM), and ethanol increased hippuric acid (HA) excretion in urine more significantly at low (less than 250 ppm) than at high atmospheric toluene concentrations. Ethanol also expressed a similar effect on p-cresol excretion as on HA, but had little effect on o-cresol. Phenobarbital treatment promoted the urinary excretion of all of the metabolites of toluene, especially after exposure to high toluene concentration. As well as HA, benzoylglucuronide (BG) and free benzoic acid were found in urine. These are the products of the side chain metabolism of toluene. Amounts of BG could be detected when the urinary excretion of free benzoic acid exceeded 5 mumol/kg/6 h, indicating that a great deal of benzoic acid is required for the formation of BG. The Michaelis constant (Km) and the maximum rate of metabolic excretion in urine during six hours exposure (Vmax) of isozymes involved in the excretion of toluene metabolites were calculated, and correlated with the subtypes of cytochrome P-450. The significance of the result was suggested in the biological monitoring of exposure to toluene.     

Quantitation of urinary o-xylene metabolites of rats and human beings by high performance liquid chromatography

Summary A high performance liquid chromatographic method for the determination of urinary o-xylene metabolites of rats, volunteers, and workers was described. In rat urine, the major metabolite, indicated by the glucuronic acid reaction, was separated with thin-layer liquid chromatography and identified as o-toluic acid glucuronide by high performance liquid chromatography. Another rather minor metabolite was demonstrated to be o-methyl hippuric acid. One of the major urinary metabolite in volunteers administered o-xylene orally was demonstrated to be o-methylhippuric acid and another minor metabolite was o-toluic acid glucuronide. In the urine of volunteers exposed to 138 ppm of o-xylene for 3 h in an artificial exposure chamber, o-methylhippuric acid was found to be the major metabolite and a minute amount of o-toluic acid glucuronide was found to be the minor metabolite. In the urine of shipbuilding workers, using a thinner containing toluene and xylenes (o-, m- and p-), hippuric acid and methylhippuric acids (o-, m- and p-) were recognized. Thus, urinary o-methylhippuric acid could be an index of o-xylene exposure of workers.     

Exposure of workers to a mixture of toluene and xylenes. I. Metabolism.

The urinary excretion of hippuric acid and methylhippuric acid was studied in workers (233 subjects; 122 men and 111 women) exposed to toluene and xylenes in combination and in non-exposed controls (281 subjects; 141 men and 140 women) recruited from the same factories or factories of the same regions. Smoking and drinking habits of the subjects were obtained by medical interviews. From each worker, one urine sample was collected at the end of a shift and analysed for hippuric and methylhippuric acids by high performance liquid chromatography. Air samples for the estimation of toluene and xylenes were collected with diffusive personal samplers. There was a linear correlation between the time weighted average exposure either to toluene or xylene isomers and the concentrations of hippuric acid or methylhippuric acid isomers in urine. Essentially no difference was found in the correlation between quantitative exposure and excretion in the three xylene isomers. Comparison of the slopes of regression lines indicated the absence of metabolic interaction between toluene and xylenes at the measured concentrations. The metabolism of toluene and xylenes was significantly reduced among smokers or drinkers compared with non-smokers and non-drinkers.     

Urinary excretion of O-cresol and hippuric acid after toluene exposure in rotogravure printing

Summary In 62 male rotogravure printers, the time-weighted average (TWA) toluene exposure during one workweek ranged from 8 to 496 mg/m³ (median 96). Post-shift urinary excretion of hippuric acid showed a poor correlation with the air toluene concentration. Level of o-cresol excretion ranged from 0.08 to 2.37 mmol/mol creatinine and was associated with the exposure (r _s = 0.57, P<0.0001), although the variation was considerable. However, this metabolite was significantly influenced by smoking habits, both in the workers (0.34 vs 0.10 mmol/mol creatinine after adjustment to zero exposure for the smokers and non-smokers, respectively; P = 0.03) and in 21 unexposed controls (0.18 vs 0.06 mmol/mot creatinine; P = 0.002). The excretion of these metabolites was followed during vacation, when the workers were unexposed. The shared one-compartment half-time was 44h (± SE 30, 82). After 2–4 weeks of vacation, the concentration of o-cresol was significantly higher for the smokers than the non-smokers (0.14 vs 0.06 mmol/mol creatinine; P = 0.02).No smoking-associated difference was found for the urinary hippuric acid concentration. However, there was an association between alcohol consumption and hippuric acid excretion (P = 0.03); no such difference was shown for o-cresol. These results demonstrate that hippuric acid excretion is unsuitable for biological monitoring of toluene exposure when the exposure level is below 200 mg/m³. Also, in spite of the favourable excretion kinetics, the impact of smoking and the large interindividual variation warrant the same conclusion for o-cresol as a means of monitoring low level exposure in an individual worker.     

Urinary levels of proteins and metabolites in workers exposed to toluene

Summary We measured urinary excretion of albumin and retinol-binding proteins to investigate the occurrence of early renal dysfunction in 45 paint workers exposed principally to toluene, and in the same number of unexposed control subjects matched individually for sex and age. Two biological indicators of personal toluene absorption, namely urine hippuric acid and o-cresol, were also measured in the exposed subjects. A significantly higher level and increased prevalence of elevated retinol-binding protein in the urine of exposed workers was found, whereas no significant difference in urinary albumin concentration was seen between the two groups. Urinary concentrations of retinol-binding protein was correlated (r = 0.399, P < 0.006) with that of o-cresol, but not with hippuric acid or employment duration. The results suggest a dose-dependent early tubular effect due to toluene exposure that might be useful for monitoring individuals exposed to toluene at work.     

Urinary levels of proteins and metabolites in workers exposed to toluene

We measured urinary excretion of albumin and retinol-binding proteins to investigate the occurrence of early renal dysfunction in 45 paint workers exposed principally to toluene, and in the same number of unexposed control subjects matched individually for sex and age. Two biological indicators of personal toluene absorption, namely urine hippuric acid and o-cresol, were also measured in the exposed subjects. A significantly higher level and increased prevalence of elevated retinol-binding protein in the urine of exposed workers was found, whereas no significant difference in urinary albumin concentration was seen between the two groups. Urinary concentrations of retinol-binding protein was correlated (r = 0.399, P less than 0.006) with that of o-cresol, but not with hippuric acid or employment duration. The results suggest a dose-dependent early tubular effect due to toluene exposure that might be useful for monitoring individuals exposed to toluene at work.     

Effect of ethanol,cimetidine and propranolol on toluene metabolism in man

Summary In a climatic exposure chamber four healthy volunteers were exposed to 100ppm toluene, 100ppm toluene + ethanol, 100ppm toluene + cimetidine, and 100ppm toluene + propranolol for 7h each at random over four consecutive days. A control experiment and 3.5 h of exposure to 200 ppm toluene were also performed. Ethanol inhibited toluene metabolism by 0.5 as expressed by the urinary excretion of two of the metabolites of toluene, namely o-cresol and hippuric acid. In agreement with this, the mean alveolar concentration of toluene was greater by 1.7 during ethanol exposure; 45 min after discontinuation of exposure the increase was by 3.3. Neither cimetidine nor propranolol changed toluene metabolism significantly. The results indicate that ethanol may prolong the time interval in which toluene is retained in the human body in persons simultaneously exposed to ethanol and toluene. When using o-cresol or hippuric acid in biological monitoring of persons occupationally exposed to toluene, the consumption of ethanol should be considered.Supported by grants from the Working Environment Fund, Denmark     

Urinary hippuric acid and orthocresol excretion in man during experimental exposure to toluene.

It is not known whether urinary excretion of hippuric acid (HA) or orthocresol (O-Cr) is to be preferred for the biological monitoring of workers with occupational exposure to toluene. To study this, 42 printing trade workers with more than 10 years' exposure to a mixture of organic solvents including toluene (0-20 ppm) and 43 control subjects matched by age, smoking habits, and living accommodation were investigated. Each matched pair was randomised to an experimental exposure of either 100 ppm or 0 ppm toluene for 6.5 hours under controlled conditions in an exposure chamber. Urinary excretion of HA and O-Cr was determined by high pressure liquid chromatography from samples obtained before exposure, during the first three hours, and during the last 3.5 hours of exposure. No difference in HA and O-Cr excretion was found between printing trade workers and controls. The median O-Cr excretion increased 29 times during exposure, whereas the HA excretion increased only five times. Thus only 3% of the O-Cr excretion originated from other sources than toluene whereas the corresponding value for HA was 19%. Standardisation of the concentrations of HA and O-Cr in relation to urinary creatinine reduced the relative variation by 29% and 56% respectively. This was not reduced further by expressing the excretions as average excretion rates based on total volume of urine collected. Background urinary O-Cr excretion was three to four times higher among smokers than non-smokers, probably due to the content of O-Cr in cigarettes. The O-Cr excretion in unexposed smokers was, however, 10 times lower that that of the non-smokers during the end of the experimental exposure to 100 ppm toluene.     

Excretion of hippuric acid and m- or p-methylhippuric acid in the urine of persons exposed to vapours of toluene and m- or p-xylene in an exposure chamber and in workshops, with specific reference to repeated exposures

Ogata, M., Takatsuka, Y., and Tomokuni, K. (1971).Brit. J. industr. Med.,28, 382-385. Excretion of hippuric acid and m- or p-methylhippuric acid in the urine of persons exposed to vapours of toluene and m- or p-xylene in an exposure chamber and in workshops, with specific reference to repeated exposures. Four male volunteers were exposed to 200 p.p.m. of toluene for five one-hour periods separated by one-hour intervals. The excretion curve of hippuric acid showed multi-peaks, and almost concided with a theoretical curve previously described. The fraction of the toluene absorbed which was accounted for as hippuric acid was only slightly lower than after a single exposure.

In a paint spraying shop exposure was measured both from the concentrations of toluene in the air by a Kitagawa detector and from the exceretion of urinary hippuric acid. The results were in general agreement, with a correlation coefficient of 0·67.

Urinary hippuric acid and methylhippuric acid were determined on urines from two workers in a shipbuilding yard who used paint thinned with toluene and xylene. The concentrations of the acids varied from day to day depending on the kind and the duration of work. From the concentrations found the mean concentrations to which the workers were exposed were calculated as a fraction of the maximum allowable concentration (M.A.C.). One worker was, on this evidence, exposed to more than the combined M.A.C. on three days out of six.

     

Chronic occupational exposure to toluene

Summary Chronic occupational exposure to toluene was studied in a factory preparing tarpaulins. Seventy-eight workers were studied; 46 were exposed to various concentrations of toluene in air (20–200 ppm), 32 were unexposed workers in the same factory. In many cases the exposure had lasted for 10–20 years. The urinary hippuric acid excretion at the end of work shift showed good correlations to toluene concentrations in air, and it seems to be a good measure of exposure. The hippuric acid in urine samples collected overnight showed that elimination of toluene still occurs several hours after exposure. Most of the biological parameters measured showed no correlation to toluene exposure. The blood leukocyte count did show slight positive correlations to toluene exposure, but even this parameter stayed inside the range of normal values. The occurrence of chronic diseases, drug using habits, and drinking and smoking habits did not show any correlations to toluene exposure.This study has been supported by the grant of Y. Jahnsson Foundation in Finland     

Urinary excretion of hippuric acid and m- or p-methylhippuric acid in the urine of persons exposed to vapours of toluene and m- or p-xylene as a test of exposure

Ogata, M., Tomokuni, K., and Takatsuka, Y. (1970).Brit. J. industr. Med.,27, 43-50. Urinary excretion of hippuric acid and m- or p-methylhippuric acid in the urine of persons exposed to vapours of toluene and m- or p-xylene as a test of exposure. Twenty-three male volunteers were exposed in groups of four or five to toluene and m- and p-xylene vapour for periods of 3 hours or of 7 hours with one break of an hour. Urine was collected at hourly intervals for several hours, and thereafter all urine was collected until 18 hours after the end of the exposure period, and was analysed for hippuric and methylhippuric acids. It was shown that hippuric acid was excreted equivalent to 68% of the toluene absorbed, and m-methylhippuric acid equivalent to 72% of the m-xylene absorbed. Up to hydrocarbon concentrations of 200 ppm the total quantity of hippuric acids excerted was proportional to the total exposure (ppm × hours). In descending order of precision the following were also related to exposure: rate of excretion during the exposure period; concentrations of hippuric acid in urine corrected to constant urine density; and concentrations in urine uncorrected for density. The last could not be used to calculate exposure, but the others could be to give screening tests to show whether workmen could have been exposed to concentrations greater than the maximum allowable.

The effects of exposure on blood pressure, pulse rate, flicker value, and reaction time were measured. There were some variations which suggested that the MAC of toluene should be set higher than 200 ppm.

     

Mutual metabolic suppression between benzene and toluene in man

Summary The exposure intensity during a shift and the metabolite levels in the shift-end urine were examined in male workers exposed to either benzene (65 subjects; the benzene group), toluene (35 subjects; the toluene group), or a mixture of both (55 subjects; the mixture group). In addition, 35 non-exposed male workers (the control group) were similarly examined for urinary metabolites to define background levels. A linear relationship was established between the intensity of solvent exposure and the corresponding urinary metabolite levels (i.e. phenol, catechol and quinol from benzene, and hippuric acid and o-cresol from toluene) in each case when one of the three exposed groups was combined with the control group for calculation. Comparison of regression lines in combination with regression analysis disclosed that urinary levels of phenol and quinol (but not catechol) were lower in the mixture group than in the benzene group when the intensities of exposure to benzene were comparable, indicating that the biotransformation of benzene to phenolic compounds (excluding catechol) in man is suppressed by co-exposure to toluene. Conversely, metabolism of toluene to hippuric acid was suppressed by benzene co-exposure. Conversion of toluene to o-cresol was also reduced by benzene, but to a lesser extent. The significance of the present findings on the mutual suppression of metabolism between benzene and toluene is discussed in relation to solvent toxicology and biological monitoring of exposure to the solvents.     

Toluene metabolism during exposure to varying concentrations combined with exercise

Summary The urinary excretion of hippuric acid (HA) and ortho-cresol (O-cr) in man was measured in two studies of 7-h exposure to toluene in a climate chamber, either constant concentration of 100 ppm or varying concentrations containing peaks of 300 ppm but with a time-weighted average of 100 ppm. In Study A, four males were exposed to clean air and to constant and varying concentrations of toluene in combination with rest and with 100 W exercise in 140 min. Exercise increased end exposure excretion rate of HA and O-cr by 47 and 114%, respectively. After exposure, all excess HA was excreted within 4 h, while O-cr was eliminated with a half life of about 3 h. Alveolar air concentration of toluene varied between 21 and 31 ppm during constant exposure and between 13 and 57 ppm during varying exposure, but no difference in mean alveolar toluene concentration or in metabolite excretion was seen between the exposure schedules. In Study B, 32 males and 39 females aged between 31 and 50 years were exposed once to either clean air, constant or varying concentrations of toluene. Background excretion rate of HA was 0.97 ± 0.75 mg/min (1.25 ± 1.05 g/g creatinine) and rose to 3.74 ± 1.40 mg/min (3.90 ± 1.85 g/g cr) during the last 3 h of exposure to 100 ppm toluene. The corresponding figures for O-cr were 0.05 ± 0.05 g/min (0.08 ± 0.14 mg/g cr), and 2.04 ± 0.84 g/min (2.05 ± 1.18 mg/g cr). The individual creatinine excretion rate was considerably influenced by sex, body weight and smoking habits, thus influencing the metabolite concentration standardised in relation to creatinine. It is concluded that both metabolites are estimates of toluene exposure. O-cr is more specific than HA, but the individual variation in excretion of both metabolites is large, and when implementing either of them as biological exposure indices, the influence of sex, body size, age as well as consumption of tobacco and alcohol has to be considered.     

Exposure to toluene increases the urinary excretion of D-glucaric acid.

Workers at a printing plant exposed to low concentrations of toluene (43-401 mg/m3, median 155 mg/m3) had increased urinary D-glucaric acid (3.55-5.12 mmol/mol creatinine) excretion at the end of the shift compared with controls (2.45-3.35 mmol/mol creatinine). No increase was found after the summer holiday (1.92-2.89 mmol/mol creatinine) but excretion had increased two weeks later (4.05-5.55 mmol/mol creatinine). These changes in the excretion of D-glucaric acid were not correlated to levels of exposure, to changes of urinary hippuric acid and o-cresol half lives (three to eight hours), nor to o-cresol/hippuric acid concentration ratios when measured at the end of daily exposure. Since a significant intra and interindividual variability of urinary D-glucaric acid was found in all groups, urinary D-glucaric acid excretion is suitable to monitor group but not individual exposure.     

Correlation of xylene exposure and methyl hippuric acid excretion in urine among paint industry workers

Different calculations of methyl hippuric acid excretion in urine were correlated to the time-weighted average (TWA) of the xylene exposure of a complete workday for 40 paint industry workers exposed to 12 different solvents. The 8-h TWA xylene exposure varied between 0 and 865 (median 69) mg/m3. The amount of methyl hippuric acid excreted in about 24 h showed only a slightly higher linear correlation to the xylene exposure than the amount of methyl hippuric acid excreted per hour during the latter part of the workshift among the 37 subjects exposed to TWA xylene air concentrations of 0-200 mg/m3. It was concluded that the methyl hippuric acid excretion rate during the latter part of the workshift can be used for crude xylene exposure categorizations.