Determination of urinary hippuric acid and o-cresol levels as biological indicators of toluene exposure in shoe-workers and glue sniffers.

In this study, groups exposed to toluene either intentionally (glue sniffers) or unintentionally (shoe-workers) were compared. The groups were evaluated in terms of urinary levels of the toluene metabolites hippuric acid and o-cresol. Results were also compared with control values. Hippuric acid levels were determined by high performance liquid chromatography and o-cresol levels by gas liquid chromatography. The levels of hippuric acid and o-cresol were found to be statistically significantly higher in glue sniffers than in shoe-workers (p <0.001) or controls (p <0.001). In addition, the differences between the levels of urinary hippuric acid and o-cresol in the shoe-workers and in the controls were statistically significant (p <0.05 and p <0.001, respectively). These results suggest that extremely high levels of urinary hippuric acid and o-cresol indicate massive exposure to toluene.     

Metabolite ratio of toluene-exposed rotogravure printing plant workers reflects individual mutagenic risk by sister chromatid exchanges

The study involved a group of 42 printing plant workers and a control group of 45 blood donors. At the working places, the ambient air-toluene concentration amounted from 141 to 328 mg/m(3). Sister chromatid exchanges (SCEs) were significantly elevated by three units in the exposed group. In this group, the concentration of urinary toluene metabolites was also considerably increased-hippuric acid was four times higher and the o-cresol and p-cresol fractions were twice as high. Results of toluene monitoring of ambient air- or blood-toluene concentrations did not show any relationships with individual SCE. While these SCE values revealed only a weak relationship with the corresponding hippuric acid data, a significant correlation with the cresols, which are known to be more genotoxic than hippuric acid, appeared in highly exposed workers. An attempt was made to consider the individual metabolic balance of toluene excretion products. For that reason individual cresol to hippuric acid ratios were calculated and related to corresponding SCE values. In all investigated subpopulations of the exposed group, this ratio correlated with SCE at a level of high significance. This strong interrelationship is a powerful argument for the genotoxic behavior of toluene. Furthermore, the individual metabolic balance, as a consequence of genetic polymorphism, should be considered in the discussion about genetic risk of toluene.     

Blood lead levels in glue sniffers

The aim of this study was to investigate blood levels of lead (Pb) among adolescents with glue sniffing in Turkey. Blood Pb levels were measured in 30 adolescent glue sniffers by atomic absorption spectrophotometry and compared with those of the 30 healthy adolescents. The Pb contents of various glue preparations marketed in Turkey and used by the abusers were also measured. Blood Pb levels were significantly higher in the study group when compared to the control group. Pb was detected at considerably high levels in the contents of all the various glue preparations most commonly used by the cases in the study group. The increased blood Pb levels in glue sniffers may be related to the high lead contents of glues marketed in Turkey. The blood Pb levels and signs of Pb toxicity should be investigated in examination of glue sniffers.     

Exhaled breath condensate as matrix for toluene detection: A preliminary study

Abstract

The study was designed to investigate whether exhaled breath condensate, obtained by cooling exhaled air in spontaneous breathing, could be a suitable matrix for toluene quantitative analyses. Nine healthy subjects were exposed for a short period (20 min) to a known concentration of toluene. Exhaled breath condensate samples were collected before and at the end of the exposure, while the environmental concentration of toluene was continuously monitored. Toluene was analysed by head-space gas-chromatography mass spectrometry, and assay repeatability was also estimated in vitro. Baseline and post-exposure measurement of hippuric acid, the urinary toluene metabolite, was performed to assess current toluene exposure. Before the exposure toluene concentrations in the exhaled breath condensate were lower than the detectable limit in all subjects, while after the exposure toluene was detectable with a median value 0.35 µg l^?1 (range 0.15–0.55 µg l^?1) in all the exhaled breath condensate samples. As compared with the standard calibration in distilled water, the curves obtained by exhaled breath condensate were linear and comparable with the range examined in vivo for toluene. A significant correlation was found between the environmental toluene levels and toluene in the exhaled breath condensate at the end of exposure. Furthermore, a significant relationship between increased exhaled breath condensate toluene levels and urinary hippuric acid after the exposure was found. In conclusion, exhaled breath condensate is a promising matrix for toluene assessment, although its application in humans requires further investigations.     

Evaluation of genotoxicity in a group of workers from a petroleum refinery aromatics plant

Petroleum refinery workers are potentially exposed to a wide range of petroleum-derived hydrocarbons and chemical substances used in the manufacturing of petroleum derivatives. Benzene, toluene and xylene (BTX) are produced by distillation in the aromatics units and used as raw materials for petrol and petrochemical products. The aim of this study was to evaluate the genotoxic effects of occupational exposure to BTX in a petroleum refinery in the North of Portugal. The exposed group consisted of 48 workers from the aromatics plant and the control group consisted of 30 persons matched for various confounding factors. Chromosome aberrations (CA), micronuclei (MN), and DNA damage (evaluated by means of the comet assay) were measured in peripheral blood leukocytes. t,t-Muconic acid (t,t-MA), hippuric acid (HA) and methylhippuric acid (MHA) concentrations were measured in urine samples collected at the end of the workshift. The results suggest that occupational exposure to toluene and xylene is very low. A statistically significant increase in t,t-MA excretion was found in the exposed group although t,t-MA levels were found to be lower than the biological exposure index (BEI). Significant increases were found for CA, MN and comet tail length (TL) in the exposed group (p<0.05). No association was found between tobacco smoking and the effect biomarkers analysed. A positive association was found between CA and MN with age in the control group (p<0.05).     

Influence of various mixtures of inhaled toluene and xylene on the biological monitoring of exposure to these solvents in rats.

The present study was undertaken to describe the influence of simultaneous exposure by inhalation to toluene and xylene on some aspects of their respective metabolic disposition. Adult male rats were exposed acutely (5 h) to 75, 150, and 225 ppm of toluene or xylene and to various mixtures of these solvents: toluene (75 ppm) and xylene (225 ppm), toluene (150 ppm) and xylene (150 ppm), toluene (225 ppm) and xylene (75 ppm). Compared with single exposure, simultaneous exposure resulted in lower amounts of excreted hippuric acid (20-30%) and methylhippuric acids (4-40%) in urine over a period of 24 h, even though significant differences were seen only with the toluene (150 ppm) and xylene (150 ppm) combination. In addition, increased concentrations of solvents in blood (toluene, 230%; xylene, 500%) and in brain (toluene, 230%; xylene, 320%) were found during the immediate post-exposure period. Simultaneous exposure also enhanced the pulmonary elimination of both solvents (toluene, 190-240%; xylene, 340-650%). Influence of repeated simultaneous exposure (9 days) was investigated for the toluene (150 ppm) and xylene (150 ppm) combination and the results compared with those of repeated exposure to each solvent administered singly. Under these conditions, repeated simultaneous exposure decreased the excretion of urinary metabolites, but only after the first exposure. On the other hand, simultaneous exposure resulted in significantly higher concentrations of toluene (210%) and xylene (240%) in blood throughout the entire 9-day exposure period. These results strongly suggest mutual metabolic interactions (inhibition) between toluene and xylene that affect the metabolic disposition of both solvents and ultimately the biological monitoring of data of exposure to a combination of solvents in rats.     

Comparison of genetic damage in Brazilian footwear-workers exposed to solvent-based or water-based adhesive

Research has shown that workers employed in footwear manufacture are at increased risk of some cancers, the strongest evidence being for nasal cancer and leukemia. Footwear-workers are routinely exposed to complex mixtures of solvents in degreasers, cleaners, primers, and adhesives used in the production process as toluene, n-hexane, acetone, and possibly dust particles, additives in shoe materials and degradation products of materials. The recognition of the potential health-hazards of solvent-based adhesives (SBAs) has lead to the development of adhesives with no organic solvents, the water-based adhesives (WBA). We investigated footwear-workers (all males) exposed to SBA (n=29) (for 3.98+/-4.13 years), and WBA (n=16), which had spent the six months previous to the study employed in an experimental section which used only water-based adhesives, although they had previously worked in sections which used solvent-based adhesives (for 5.80+/-4.03 years); 25 healthy subjects were used as controls. The Comet assay and the micronucleus test were used as endpoints, while the traditional parameters for assessing exposure to toluene in organic mixtures by measuring the concentration of urinary hippuric acid were also assessed. Our results showed a significantly lower mean concentration of hippuric acid in the control group than found in the SBA (P<0.001) and WBA (P<0.05) groups. The Comet assay results showed that there was a significant increase in the mean damage index for the SBA (P<0.001) group in comparison to the WBA group and control (P<0.05). For the micronucleus test in binucleated lymphocytes and exfoliated buccal cell, the three groups were not statistically different. Our study demonstrated that water-based adhesives are clearly a better option for safeguarding the health of footwear-workers, even with possibility of isocyanate presence, while the positive results observed in SBA group might be explained by chloroprene presence in the adhesive.     

Cytogenetic analysis of buccal cells from shoe-workers and pathology and anatomy laboratory workers exposed to n-hexane, toluene, methyl ethyl ketone and formaldehyde.

People employed in the shoe manufacture and repair industry are at an increased risk for cancer, the strongest evidence being for nasal cancer and leukaemia. A possible causal role for formaldehyde is likely for cancer of the buccal cavity and nasopharynx. Exfoliated buccal cells are good source of tissue for monitoring human exposure to inhaled and ingested occupational and environmental genotoxicants. To assess the cytogenetic damage related to occupational exposure to airborne chemicals during shoe-making and the processes in pathology and anatomy laboratories, the micronuclei (MN) count per 3000 cells was measured in buccal smears from shoe-workers (group I, n = 22) exposed to mainly n-hexane, toluene and methyl ethyl ketone (MEK) and from anatomy and pathology staff (group II, n = 28) exposed to formaldehyde (FA). Eighteen male university staff were used as controls. The mean time-weighted average (TWA) concentrations of n-hexane, toluene and MEK in 10 small shoe workshops were 58.07 p.p.m., 26.62 p.p.m. and 11.39 p.p.m., respectively. The measured air concentrations of FA in the breathing zone of the anatomy and pathology laboratory workers were between 2 and 4 p.p.m. Levels of 2,5-hexadione (2,5-HD) and hippuric acid (HA), metabolic markers of n-hexane and toluene exposure, respectively, were significantly higher in the urine of workers in group I than in control subjects (p < 0.001 and p < 0.01, respectively). The mean (+/- SD) MN (0/00) [corrected] frequencies in buccal mucosa cells from workers in group I, group II and controls were 0.62 +/- 0.45%, 0.71 +/- 0.56% and 0.33 +/- 0.30%, respectively (p < 0.05 and p < 0.05 compared with controls for group I and group II, respectively). The effects of smoking, age and duration of exposure on the frequency of micronucleated buccal cells from workers in all three groups studied were also evaluated. Overall, the results suggest that occupational exposure to organic solvents, mainly n-hexane, toluene, MEK and FA, may cause cytogenetic damage in buccal cells and that use of exfoliated buccal cells seems to be appropriate to measure exposure to organic solvents.     

The fate of benzoic acid in various species

1. The urinary excretion of orally administered [¹⁴C]benzoic acid in man and 20 other species of animal was examined. 2. At a dose of 50mg/kg, benzoic acid was excreted by the rodents (rat, mouse, guinea pig, golden hamster, steppe lemming and gerbil), the rabbit, the cat and the capuchin monkey almost entirely as hippuric acid (95–100% of 24h excretion). 3. In man at a dose of 1mg/kg and the rhesus monkey at 20mg/kg benzoic acid was excreted entirely as hippuric acid. 4. At 50mg/kg benzoic acid was excreted as hippuric acid to the extent of about 80% of the 24h excretion in the squirrel monkey, pig, dog, ferret, hedgehog and pigeon, the other 20% being found as benzoyl glucuronide and benzoic acid, the latter possibly arising by decomposition of the former. 5. On increasing the dose of benzoic acid to 200mg/kg in the ferret, the proportion of benzoyl glucuronide excreted increased and that of hippuric acid decreased. This did not occur in the rabbit, which excreted 200mg/kg almost entirely as hippuric acid. It appears that the hedgehog and ferret are like the dog in respect to their metabolism of benzoic acid. 6. The Indian fruit bat produced only traces of hippuric acid and possibly has a defect in the glycine conjugation of benzoic acid. The main metabolite in this animal (dose 50mg/kg) was benzoyl glucuronide. 7. The chicken, side-necked turtle and gecko converted benzoic acid mainly into ornithuric acid, but all three species also excreted smaller amounts of hippuric acid.     

Exhaled breath condensate as matrix for toluene detection: A preliminary study

The study was designed to investigate whether exhaled breath condensate, obtained by cooling exhaled air in spontaneous breathing, could be a suitable matrix for toluene quantitative analyses. Nine healthy subjects were exposed for a short period (20 min) to a known concentration of toluene. Exhaled breath condensate samples were collected before and at the end of the exposure, while the environmental concentration of toluene was continuously monitored. Toluene was analysed by head-space gas-chromatography mass spectrometry, and assay repeatability was also estimated in vitro. Baseline and post-exposure measurement of hippuric acid, the urinary toluene metabolite, was performed to assess current toluene exposure. Before the exposure toluene concentrations in the exhaled breath condensate were lower than the detectable limit in all subjects, while after the exposure toluene was detectable with a median value 0.35 µg l^-1 (range 0.15-0.55 µg l^-1) in all the exhaled breath condensate samples. As compared with the standard calibration in distilled water, the curves obtained by exhaled breath condensate were linear and comparable with the range examined in vivo for toluene. A significant correlation was found between the environmental toluene levels and toluene in the exhaled breath condensate at the end of exposure. Furthermore, a significant relationship between increased exhaled breath condensate toluene levels and urinary hippuric acid after the exposure was found. In conclusion, exhaled breath condensate is a promising matrix for toluene assessment, although its application in humans requires further investigations.     

Evaluation of DNA damage by the Comet assay in shoe workers exposed to toluene and other organic solvents

The alkaline single-cell gel electrophoresis (or Comet) assay was applied to evaluate DNA damage in cryopreserved peripheral blood mononuclear leukocytes from 34 female shoe workers exposed to organic solvents and a group of 19 non-exposed women. We also investigated whether the polymorphisms of glutathione S-transferase M1 (GSTM1) and T1 (GSTT1) genes affect individual level of DNA damage possibly induced by the solvent exposure. Chemical measurements of workplace air in the two factories studied showed that the workers were exposed to acetone, gasoline, and toluene in both factories and to ethylacetate and diisocyanate in one factory. In the exposed workers, the average level of blood hemoglobin was lower and that of urinary hippuric acid higher than in the non-exposed individuals. However, the occupational exposure to organic solvents did not affect the Comet values. Neither did age, smoking, or the GSTM1 genotype have any effect on the outcome of this assay. The low prevalence of the GSTT1-null genotype precluded conclusions on the influence of GSTT1 polymorphism.     

Biodegradation of ortho-cresol by a mixed culture of nitrate-reducing bacteria growing on toluene.

A mixed culture of nitrate-reducing bacteria degraded o-cresol in the presence of toulene as a primary growth substrate. No degradation of o-cresol was observed in the absence of toluene or when the culture grew on p-cresol and 2,4-dimethylphenol. In batch cultures, the degradation of o-cresol started after toluene was degraded to below 0.5 to 1.0 mg/liter but continued only for about 3 to 5 days after the depletion of toluene since the culture had a limited capacity for o-cresol degradation once toluene was depleted. The total amount of o-cresol degraded was proportional to the amount of toluene metabolized, with an average yield of 0.47 mg of o-cresol degraded per mg of toluene metabolized. Experiments with [ring-U-14C]o-cresol indicated that about 73% of the carbon from degraded o-cresol was mineralized to CO2 and about 23% was assimilated into biomass after the transient accumulation of unidentified water-soluble intermediates. A mathematical model based on a simplified Monod equation is used to describe the kinetics of o-cresol degradation. In this model, the biomass activity toward o-cresol is assumed to decay according to first-order kinetics once toluene is depleted. On the basis of nonlinear regression of the data, the maximum specific rate of o-cresol degradation was estimated to be 0.4 mg of o-cresol per mg of biomass protein per h, and the first-order decay constant for o-cresol-degrading biomass activity was estimated to be 0.15 h-1.(ABSTRACT TRUNCATED AT 250 WORDS)     

Cytogenetic analysis of buccal cells from shoeworkers and pathology and anatomy laboratory workers exposed to n-hexane,toluene, methyl ethyl ketone and formaldehyde

People employed in the shoe manufacture and repair industry are at an increased risk for cancer, the strongest evidence being for nasal cancer and leukaemia. A possible causal role for formaldehyde is likely for cancer of the buccal cavity and nasopharynx. Exfoliated buccal cells are good source of tissue for monitoring human exposure to inhaled and ingested occupational and environmental genotoxicants. To assess the cytogenetic damage related to occupational exposure to airborne chemicals during shoe-making and the processes in pathology and anatomy laboratories, the micronuclei (MN) count per 3000 cells was measured in buccal smears from shoe-workers (group I, n = 22) exposed to mainly n-hexane, toluene and methyl ethyl ketone (MEK) and from anatomy and pathology staff (group II, n = 28) exposed to formaldehyde (FA). Eighteen male university staff were used as controls. The mean time-weighted average (TWA) concentrations of n-hexane, toluene and MEK in 10 small shoe workshops were 58.07 p.p.m., 26.62 p.p.m. and 11.39 p.p.m., respectively. The measured air concentrations of FA in the breathing zone of the anatomy and pathology laboratory workers were between 2 and 4 p.p.m. Levels of 2,5-hexadione (2,5-HD) and hippuric acid (HA), metabolic markers of n-hexane and toluene exposure, respectively, were significantly higher in the urine of workers in group I than in control subjects (p < 0.001 and p < 0.01, respectively). The mean (±SD) MN frequencies in buccal mucosa cells from workers in group I, group II and controls were 0.62±0.45%, 0.71±0.56% and 0.33±0.30%, respectively (p < 0.05 and p < 0.05 compared with controls for group I and group II, respectively). The effects of smoking, age and duration of exposure on the frequency of micronucleated buccal cells from workers in all three groups studied were also evaluated. Overall, the results suggest that occupational exposure to organic solvents, mainly n-hexane, toluene, MEK and FA, may cause cytogenetic damage in buccal cells and that use of exfoliated buccal cells seems to be appropriate to measure exposure to organic solvents.     

Mutants of Pseudomonas cepacia G4 defective in catabolism of aromatic compounds and trichloroethylene

Pseudomonas cepacia G4 possesses a novel pathway of toluene catabolism that is shown to be responsible for the degradation of trichloroethylene (TCE). This pathway involves conversion of toluene via o-cresol to 3-methylcatechol. In order to determine the enzyme of toluene degradation that is responsible for TCE degradation, chemically induced mutants, blocked in the toluene ortho-monooxygenase (TOM) pathway of G4, were examined. Mutants of the phenotypic class designated TOM A- were all defective in their ability to oxidize toluene, o-cresol, m-cresol, and phenol, suggesting that a single enzyme is responsible for conversion of these compounds to their hydroxylated products (3-methylcatechol from toluene, o-cresol, and m-cresol and catechol from phenol) in the wild type. Mutants of this class did not degrade TCE. Two other mutant classes which were blocked in toluene catabolism, TOM B-, which lacked catechol-2,3-dioxygenase, and TOM C-, which lacked 2-hydroxy-6-oxoheptadienoic acid hydrolase activity, were fully capable of TCE degradation. Therefore, TCE degradation is directly associated with the monooxygenation capability responsible for toluene, cresol, and phenol hydroxylation.     

Progression of addiction careers in young adult solvent misusers.

Four case histories of a representative sample of young adult glue sniffers who had progressed to using illicit drugs or abusing alcohol or both contained several common factors that may have been relevant in the transition. For example, all had suffered parental deprivation or rejection in childhood leading to a state of chronic stress, and all had progressed from sniffing glue to inhaling heroin and thence to "snorting" and injecting heroin. The exact reason for the transition from glue sniffing to misusing other substances was not clear. Nevertheless, although these cases represent only a minority of glue sniffers, the problem of transition may be more common than hitherto supposed.     

Benzoic acid metabolism in presence of Schistosoma mansoni infection in mice

The metabolism of benzoic acid was examined in S. mansoni infected CBA mouse. The result showed that control animals dosed with 150 mg/kg benzoic acid resulted in urinary excretion of two metabolites, hippuric acid and benzoic acid glucuronide. Administration of the same dose to animal carrying S. mansoni for a period of over 6 weeks resulted in decreased formation of hippuric acid and total elimination of benzoic acid by glucuronide pathway.     

Toluene inhalation-induced adrenocortical hypertrophy and endocrinological changes in rat

Rats were exposed to toluene (1,500 ppm for 4 hr per day) for 7 days. The body weight of the rats was significantly lower and the weight of the adrenal gland was significantly higher in the toluene inhalation group compared to the controls. Microscopically, there was no obvious change in the medulla, but hypertrophy of the cortex was observed in the toluene inhalation group. And, the size of adrenocortical cells in treated-rats was also significantly enlarged than the control. Immunohistochemical staining did not show a clear difference in localization of aldosterone-positive cells between the control and inhalation groups. Expansion of the corticosterone-positive area consistent with the cortical hypertrophy was recognized in the inhalation group. Enhancement of 72 kD-heat-shock protein (HSP70)-expression in the toluene inhalation group was not observed. Neither stress nor damage to cortical cells due directly to toluene exposure was observed in the cortex. Also, there was no obvious difference in the anti-proliferating cell nucleus antigen (PCNA)-immunostaining between control and inhalation groups. Thus, it is suspected that cortical hypertrophy was the result of cell enlargement due to the stimulation of the cortical cells. Corticotropin-releasing factor (CRF) immunoreactivity in the paraventricular nucleus (PVN) was increased in the inhalation group. Concentration of plasma ACTH was elevated significantly by toluene exposure. The amounts of mRNA of adrenocortical steroid metabolism gene, cytochrome side-chain cleavage (P450scc), was also increased by toluene inhalation. Toluene exposure might induce adrenocortical hypertrophy via the hypothalamus-pituitary-adrenal gland (HPA) axis.     

The effect of infusion of hypertonic saline on glomerular filtration rate and arginine vasotocin, prolactin and aldosterone in the domestic chicken

Domestic fowl were infused for 60 min with isotonic saline followed by 90 min with hypertonic saline. Plasma electrolyte concentrations, osmolality and haematocrit were measured. Urine electrolyte excretion rates, osmolar output and urine flow rates were also monitored. From these results fractional excretions of electrolytes were calculated. The renal function markers inulin and ρ-amino hippuric acid were infused to enable the measurement of glomerular filtration rate and plasma clearance of ρ-amino hippuric acid, respectively. Plasma samples were also taken to assay for the hormones prolactin, aldosterone and arginine vasotocin. Plasma electrolytes and osmolality, fractional excretion of electrolytes and osmolar output all increased, while haematocrit decreased, throughout the experiment. However, no significant change was found in urine flow rate and little change was seen in glomerular filtration rate. The clearance of ρ-amino hippuric acid, which provides an indication of renal plasma flow, increased during hypertonic saline infusion. Plasma concentrations of aldosterone and prolactin decreased during the experiment and plasma concentrations of arginine vasotocin increased. Infusion of hypertonic saline had no consistent effect on glomerular filtration rate, which may be due to conflicting influences of expansion of the extracellular fluid volume and increased plasma osmolality. Accepted: 19 January 1998     

Evaluation of genotoxicity and oxidative damage in painters exposed to low levels of toluene

Toluene is an organic solvent used in numerous processes and products, including industrial paints. Toluene neurotoxicity and reproductive toxicity are well recognized; however, its genotoxicity is still under discussion, and toluene is not classified as a carcinogenic solvent. Using the comet assay and the micronucleus test for detection of possible genotoxic effects of toluene, we monitored industrial painters from Rio Grande do Sul, Brazil. The putative involvement of oxidative stress in genetic damage and the influences of age, smoking, alcohol consumption, and exposure time were also assessed. Although all biomarkers of toluene exposure were below the biological exposure limits, painters presented significantly higher DNA damage (comet assay) than the control group; however, in the micronucleus assay, no significant difference was observed. Painters also showed alterations in hepatic enzymes and albumin levels, as well as oxidative damage, suggesting the involvement of oxidative stress. According to multiple linear regression analysis, blood toluene levels may account for the increased DNA damage in painters. In summary, this study showed that low levels of toluene exposure can cause genetic damage, and this is related to oxidative stress, age, and time of exposure.     

Effect of hippuric acid on the gaschromatographic retention of S-phenylmercapturic acid

S-phenylmercapturic acid (PMA) is one specific urinary biomarker of low-level benzene exposure. It is used for biological monitoring of benzene-exposed workers in the petrochemical industry and normally ranges from non-measurable to 10 microg/l levels in non-exposed non-smoking subjects. Benzene-exposure caused by workplace or lifestyle sources is frequently accompanied by toluene exposure, which can cause the occurrence of high levels (from 10 mg/l to more than 2000 mg/l) of hippuric acid (HA) in urine. Both solvents are toxic, and benzene is classified as a human carcinogen. The biological monitoring of benzene and toluene is therefore required for preventive care of exposed workers health. In this study a GC-MS method was adopted for measuring urinary PMA, which involved liquid-liquid extraction (LLE) with ethyl acetate from acidified urine and esterification with 0.5 N hydrochloric acid in methanol. The method evidenced a GC effect in a conventional HP-5 (30 m x 0.25 mm i.d., 0.25 microm film-thickness) methyl-phenylsilicone capillary column produced by HA on PMA. The results demonstrate that HA at concentrations as low as 250 mg/l can delay the elution of PMA and labelled internal standard from the column. The recognition and discussion of this particular GC phase soaking effect may be of help for those who are occupied in the determination of PMA and of urinary acidic metabolites by GC.