Association of Lung Function With Benzene, Toluene and Xylenes (Btx) in End-exhaled Air in Gas Station Attendants

doi:10.21203/rs.3.rs-1528048/v1

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Association of Lung Function With Benzene, Toluene and Xylenes (Btx) in End-exhaled Air in Gas Station Attendants

https://doi.org/10.21203/rs.3.rs-1528048/v1

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Background - Gas station attendants are exposed to benzene, toluene and xylenes (BTX) from fuels. The aim was to study the association between this exposure and spirometry parameters in gas station attendants and office workers as controls.

Method – In 44 male gas station attendants and 38 office workers in the Gampaha district of Sri Lanka spirometry was performed according to the guidelines of the American Thoracic Society (ATS). Pre- and post-shift end-exhaled air samples were collected from a subset of both study groups and analysed for BTX by gas chromatography-mass spectrometry.

Results – Among gas station attendants (n = 20) median pre-/post-shift exhaled air concentrations (ng/L) were: benzene 11.23/21.05; toluene 10.39/22.19; m/p-xylene 1.62/2.45; o-xylene 1.01/1.45. For controls (n = 11) these values were (ng/L): 9.47/12.04, 3.13/4.00, 1.52/1.25 and 0.50/0.48, respectively. The peak expiratory flow rate (PEFR) and %-predicted PEFR were significantly lower among gas station attendants compared to controls (p < 0.012). The % predicted FVC and FEV₁/FVC ratio were not different but in gas station attendants benzene in post-shift samples was associated with %-predicted FVC (Spearman’s correlation coefficient of -0.469, p = 0.037). In addition, the %-predicted FVC was lower in gas station attendants working ≥ 5 years compared to those who worked < 5 years. In controls a distinct pattern was observed, indicating a moderate to strong association of post-shift toluene and m/p-xylenes with all lung function parameter but not benzene. These results were adjusted for smoking. Air pollutants other than BTX may have contributed to explaining our findings.

Conclusion – Overall, we did not observe changes to the lung function of clinical relevance that can be explained by occupational exposure to BTX. In gas station attendants we observed obstructive responses in spirometry that were most pronounced for the level of work-related benzene exposure and also associated with duration of employment. In office workers (as controls) a distinct pattern of associations with BTX exposure was observed, indicating a moderate to strong association of the exposure level of toluene and m/p-xylenes with all lung function parameters.

volatile organic compounds

gasoline

road traffic

biomonitoring

spirometry

Increased intensity of road traffic leads to higher levels of air pollution, specifically in urban areas. Such airborne contaminants include nitrogen oxides (NO_x), carbon monoxide (CO), carbon dioxide (CO₂), ozone (O₃), sulphur dioxide (SO₂), volatile organic compounds (VOC) and particulate matters (PM) which are all emitted from automobile exhaust ¹. Benzene, toluene, p-, m-, o-xylene (BTX) represent toxic VOC primarily derived from gasoline fuel and exhaust fumes are related to the propulsion of passenger cars, vans, and motorcycles, representing the most important sources in road traffic.

Exposure to such pollutants results in respiratory symptoms ² and increased susceptibility to respiratory tract infections ^3,4. Pulmonary restriction and airway obstruction with involvement of small airways have been previously reported ^1,4,5.

Gas station attendants have an increased risk of exposure to fuel vapour by inhalation ⁶. Gas station attendants and workers in petrochemical industries in developing countries are particularly at risk for these deleterious effects because of the lack of adequate regulatory and protective mechanisms ³. The proximity of fuel stations to busy roads causes workers to be exposed to air pollutants other than fuel vapours as well. Lungs and airways are a primary target for adverse effects for inhaled air contaminants ¹. The average duration of daily exposure exceeds about 10h/day, especially if the employers provide overnight accommodation at the worksite. The duration of exposure may vary depending on their occupation tenure ⁷.

The health effects of exposure to fuel pollutants have not been assessed previously in Sri Lanka. This study assessed lung function using spirometry, in gas station attendants, following long term exposure to BTX, and the association between spirometry values and BTX levels in end-exhaled air, by comparing parameters between gas station attendants and office workers with a lower exposure. In addition, individual BTX exposure data were correlated with spirometry findings in a subgroup of gas station attendants and controls.

The aim was to study the association between BTX exposure and spirometry parameters in gas station attendants and office workers as controls.

Study area

Sri Lanka is an island located in South East Asia, a developing nation with a GDP of 4,310 USD per capita ⁸. The Ceylon Petroleum Corporation (CPC), the nation's oil provider, is regulated by the Ministry of Petroleum Resources Development. The nation consumes close to 98,000 barrels of crude oil per day ⁹. With over 6.6 million vehicles on its roads ¹⁰, the fuel to drive these vehicles is provided by the petrol stations employing hundreds of workers across the country. The study was carried out in the Gampaha District of the Western province of Sri Lanka from January 2018 - March 2019.

Study population

Fifty healthy male fuel attendants/handlers aged 20 to 65 years, employed for more than one year at fuel stations of the Gampaha District in the western province of Sri Lanka, were recruited. The gas station attendants were from 7 fuel stations. The control population comprised of 50 healthy age-matched male white-collar workers residing within the Ragama Grama-Niladhari (GN) division of the Gampaha District, a suburban environment North-West of Colombo with background exposure to air pollutants.

The duration of a shift, duration of employment, smoking status, presence of any health problems, symptoms of endocrine abnormalities and exposure to chemicals or fumes were recorded on a data sheet together with demographic characteristics (age, race, marital status, number of children, income). A physical examination was carried out, including measurement of height and weight. None of the study participants had a chest deformity.

End-exhaled air sampling

Exposure to BTX was studied in a subgroup of 29 gas station attendants and 16 controls. The method of exposure assessment by breath analyses as a method of human biomonitoring has been previously described ¹¹. Briefly, each participant provided an end-exhaled air sample to a BioVOC sampler (Markes, Llantrisant, UK), at the beginning of the shift (pre-shift) and the end of the shift (post-shift). To prevent contaminants from interfering with our analysis, all analytical thermal desorption (ATD) tubes were pre-cleaned by purging with helium 5.0 for four hours at 250˚C. Each study participant exhaled completely into the BioVOC sampler through a cardboard mouthpiece. Immediately after the sample was collected, the components of interest were transferred to the ATD tube. ATD tubes were capped with brass compression caps, packed in a gastight container and stored at ambient temperature (Scheepers et al., 2014).

Analysis of end-exhaled air samples

The analytical approach for determining BTX concentrations has been described by Scheepers and co-workers ¹¹. Briefly, a standard solution of BTX in methanol was used for calibration (CRM47993, Supelco/Sigma-Aldrich, Zwijndrecht, the Netherlands). Standards were prepared in a range of 0 to 100 ng/µl. Standards (n = 8 in duplicate) and field blanks (n = 2) were loaded on tubes shipped from the Netherlands to Sri Lanka and back to determine if any contamination had occurred during shipment. The ATD-tubes were analysed using thermal desorption-gas chromatography-mass spectrometry (TD-GC-MS). This system consists of an autosampler and a thermal desorption unit (series 2 Ultra and Unity, Markes International, Llantrisant, UK) online coupled to a GC-MS (Thermo Focus GC and ISQ MS, ThermoFisher Scientific, Breda, the Netherlands). We were not able to separate m- and p-xylene, and hence, results were reported as m/p-xylene. Exhaled air concentrations were expressed in ng/L by adjustment for the Bio-VOC air volume with an LOD of 0.1 ng/L.

Assessment of lung function

Spirometry was performed using a Vitalograph Alpha Touch spirometer (Vitalograph Ltd. UK) according to ATS guidelines ¹² in all study participants. The machine was calibrated before each test using a 3L calibration syringe. During spirometry, disposable bacterial filters were used to prevent cross-infection.

Spirometry was performed from January until August 2018 between 9.00 am to 11.00 am every day to minimise the biological circadian rhythm variations. All spirometry measurements were performed while they were off duty, within 2 weeks of exposure assessment, in the standing position. The manoeuvre was demonstrated to the patient by a trained investigator. Each manoeuvre was monitored by flow-volume loops and manual observation to ensure that the effort was maximal, smooth and artefact free.

Forced Vital Capacity (FVC), Forced Expired Volume in the first second (FEV₁), Peak Expiratory Flow Rate (PEFR) and Forced mid expiratory flow (FEF25-75%) were recorded. The attempt with the highest recorded parameters was used for analysis. Multiple manoeuvres (up to a maximum of 8) were performed until three satisfactory and reproducible recordings were made. The subjects were asked to rest between manoeuvres.

Repeatability was deemed satisfactory when the difference between the largest and the next largest FVC and FEV₁ was ≤ 0.150 L. The largest FVC and the largest FEV₁ observed from all of the acceptable values was used for analysis. The predicted values were calculated using published reference norms for Sri Lankan Sinhalese males ¹³. A restriction was considered clinically relevant when FVC < 80% of predicted and an obstructive ventilatory defect when the FEV₁/FVC ratio < 70%.

Statistical analysis

Because the spirometry parameters were normally distributed as confirmed by the Kolmogorov-Smirnov test (p > 0.05), the mean and SD were used as descriptive statistics. Non-parametric methods were used for subgroup analysis: a Spearman correlation coefficient was used to study associations between spirometry values and the BTX exposure levels and the partial correlation coefficient was used to control for the effect of smoking, Mann Whitney U (MWU) statistic was used to compare differences in non-normally distributed variables. The Fisher exact test was used to analyse categorical variables between the two groups. Wilcoxon signed-rank test was used to compare the paired exposure values (at the beginning and the end of the shift) of each individual. Five years of exposure as a gas station attendant was deemed adequate time for structural changes in the respiratory system to be established, and comparisons in duration of exposure by number of years of employment (5 of more years versus less than 5 years). Results were adjusted for age, height, duration of employment and smoking in regression analysis. RStudio v3.2.2 was used to develop the correlation matrix.

Complete spirometry tracings were available from 44 gas station attendants and 38 controls. The other participants had difficulty performing the manoeuvres to get three reproducible recordings on the day of testing. BTX exposure data were randomly collected from a cohort of the participants, and of the 44 participants with complete spirometry results end-exhaled air concentrations of BTX were available from 20 gas station attendants and 11 controls. These gas station attendants and controls were comparable in their physical characteristics of weight, height, and BMI. Age and height, which both influence spirometry parameters, were comparable in the two groups. A significantly higher number of gas station attendants reported waking up with a cough than the controls (p = 0.026) (Table 1).

Table 1

Demographic data and reported respiratory complaints among the cases and controls.
		Gas station attendants (n = 44)	Control (n = 38)	Significance (p)
Age (years) Mean ± SD		39.63 (12.25)	36.42 (11.59)	0.228
Ethnicity	Sinhala	43 (97%)	36 (94.73%)	0.550
Ethnicity	Other	1 (2.27%)	2 (5.27%)	0.550
Height (cm) Mean ± SD		166.00 (6.74)	167.05 (5.26)	0.439
Body weight (kg) Mean ± SD		67.78 (14.13)	65.32 (9.26)	0.362
BMI Mean ± SD		24.56 (4.51)	23.38 (2.94)	0.172
Smokers		32 (72%)	22 (42.1%)	0.158
Cough		14 (31.81%)	7 (18.42%)	0.189
Shortness of breath (SOB) at rest		5 (11.36%)	2 (5.25%)	0.363
SOB on walking		4 (9.09%)	2 (5.25%)	0.554
SOB during cycling		9 (20.45%)	3 (7.89%)	0.120
Wheezing		12 (27.27%)	8 (21.05%)	0.569
SOB when wheezing		11 (25%)	5 (13.15%)	0.813
Waking up with chest tightness		6 (13.63%)	2 (5.26%)	0.082
Waking up with SOB		3 (6.81%)	2 (5.26%)	0.550
Waking up with a cough		10 (22.72%)	4 (10.52%)	0.026
Persons on treatment for bronchial asthma		3 (6.81%)	3 (7.89%)	0.837

Among the gas station attendants, the mean (SD) number of working days per week was 4.75 (1.33), with a mean of 11.45 (2.52) hours of work per day. The exposure to fuel in the occupational setting for the gas station attendants per week was 53.14 (15.82) hours per week.

Toluene and o-xylene values in the pre-shift samples of gas station attendants were higher than in controls (p < 0.001). Post-shift values of each of the BTX components, separately, were also significantly higher among gas station attendants compared to controls (p < 0.001) (Table 2).

Table 2

Comparison of Median (IQ range) BTX concentrations of BTX (ng/L) in end-exhaled air between gas station attendants and controls in ng/L.
Substance	Sample	Exposed (n = 20) Median (IQ range)	p²	Control (n = 11) Median (IQ range)	p²	p³
Benzene	Pre-shift	11.23 (9.64–16.62)	< 0.0001	9.47 (8.73–11.20)	0.286	0.069
	Post-shift	21.05 (16.28–28.47)	< 0.0001	12.04 (8.78–13.42)	0.286	< 0.001
	Δ¹	7.95 (3.22–13.76)		1.90 (-1.66-5.24)		0.003
Toluene	Pre-shift	10.39 (8.21–21.16)	0.001	3.13 (2.83–4.93)	0.424	< 0.001
	Post-shift	22.19 (15.32–40.33)	0.001	4.00 (2.62–6.19)	0.424	< 0.001
	Δ¹	11.43 (4.10–17.00)		0.87 (-1.11-1.65)		0.001
m/p-Xylene	Pre-shift	1.62 (1.34–2.57)	0.002	1.52 (0.91–2.56)	0.091	0.536
	Post-shift	2.45 (1.97–4.22)	0.002	1.25 (1.14–1.56)	0.091	< 0.001
	Δ¹	0.97 (0.36–1.61)		0.64 (-0.45-1.07)		0.338
o-Xylene	Pre-shift	1.01 (0.82–1.47)	0.010	0.50 (0.34–0.58)	0.859	< 0.001
	Post-shift	1.45 (1.11–2.43)	0.010	0.49 (0.39–0.56)	0.859	< 0.001
	Δ¹	0.54 (0.13-1.00)		0.045 (-0.15-0.08)		0.001
¹Post-shift value minus pre-shift value; ²Pre-shift vs post-shift values in Wilcoxon signed-rank test; ³Gas station attendants vs controls in Mann-Whitney U test.

Table 3

Mean (SD) spirometry values of gas station attendants and controls.
Spirometry	Gas station attendants (n = 44)	Control (n = 38)	Significance* (p)
FVC [L]	3.27 (0.69)	3.42 (0.79)	0.346
% predicted FVC [%]	95.84 (13.57)	97.63 (18.55)	0.617
FEV₁ [L/min]	2.63 (0.60)	2.70 (0.72)	0.646
% predicted FEV₁ [%]	94.29 (12.59)	83.65 (15.62)	0.728
FEV₁/FVC ratio	79.61 (14.58)	92.66 (20.31)	0.676
PEFR [L/min]	427.39 (100.41)	488.97 (116.32)	0.012
% predicted PEFR	77.52 (17.48)	87.31 (19.38)	0.019
FEF25-75% [L/s]	2.65 (0.94)	2.71 (1.12)	0.827
% predicted FEF25-75%	84.08 (27.79)	80.50 (31.55)	0.527
* independent sample t test was used to assess the significance.

Although overall spirometry parameters were lower in gas station attendants, there was no significant difference in the mean spirometry measurements between the two groups except for PEFR and % predicted PEFR which were significantly lower among the gas station attendants compared to controls (Table 3).

Education regarding health hazards of occupation exposure was reported by 25% of gas station attendants. Use of jacket (65.9%) and boots (20.45%) were the commonest modes of personal protective measures used by the gas station attendants. Self-reported use of disposable masks was 16% and gloves was 2%. (Table S1).

We did not observe significant differences in spirometry parameters among those using protective masks compared to non-users (p > 0.05).

Smokers of both groups had lower spirometry values, but the difference was not significant (p > 0.05) (Fig. 1, Table S2). Among non-smokers (n = 28), the fuel handlers had a significantly lower PEFR compared to controls (MWU = 55.00, p = 0.016). Smokers of both groups demonstrated more obstructive and restrictive patterns on spirometry assessment (Table S3).

FVC and FEV₁ were negatively correlated with age and number of exposure days (calculated as number of years of employment x 52 weeks/year x workdays/week), also when adjusted for smoking status (using partial correlation) (Table S4). PEFR was significantly negatively correlated with age (Table S4). Age (negatively) and height (positively) correlated with spirometry parameters among controls (Table S5).

Among those who were employed as gas station attendants for five or more years, mean % predicted FVC (101.79% vs 92.45%; MWU = 133.00, p = 0.026) was statistically significantly reduced compared to those with less than 5 years of exposure (Fig. 2). However, the mean % predicted FVC was above normal limits even after 5 years of exposure. Abnormal spirometry patterns (obstruction and restriction) were not significantly higher among the group employed for 5 years or more as gas station attendant compared to those less than 5 years (restrictive pattern p = 0.099, obstructive pattern p = 0.355).

Figure 3 shows a heat map of correlations observed between exposure and outcome parameters. The weak negative and positive associations should be interpreted with caution as this may represent ‘noise’. The components of BTX exposure indicate strong associations that suggest a common source.

Among the gas station attendants there is a consistent negative correlation in the % predicted FVC across all expressions of BTX exposure. FVC, FEV₁, PEFR and FEF25-57% were negatively correlated with pre-shift and post-shift exposure (Fig. 3). These associations were more pronounced for benzene than for toluene and xylenes. The % predicted FVC was significantly negatively correlated with the pre-shift toluene (r=-0.525, p = 0.018), pre-shift o-xylene (r=-0.523, p = 0.018) and post-shift benzene (r=-0.469, p = 0.037). The percentage predicted FEV₁ significantly negatively correlated with the pre-shift toluene (r=-0.504, p = 0.024) and post-shift benzene (r=-0.493, p = 0.027). The percentage predicted FEF25-75% significantly negatively correlated with the pre-shift benzene (r=-0.695, p = 0.001), pre-shift m/p-xylene (r=-0.466, p = 0.038) and post-shift benzene (r=-0.489, p = 0.029).

A remarkable deviation is observed for the increase from pre-shift to post-shift (delta) of benzene that may be explained by divergent kinetics of benzene compared to toluene and xylenes. The results for benzene show a consistent negative correlation with lung function parameters though indicated by only moderate negative correlation coefficients. However, the measured lung function parameters remain within normal limits.

The heat map of the results observed in the office workers in Fig. 4 shows a distinct pattern from Fig. 3 for both exposure and outcome parameters. The lung function parameters in controls show a much more consist pattern of positive correlations with post-shift toluene and m/p-xylene compared to gas station attendants.

Associations of spirometry parameters with different expressions of benzene exposure are less pronounced compared to those of toluene and p/m-xylene. The weak correlations in the delta parameters indicate that there is no strong indication of an effect on lung function parameters explained from exposures during work hours. Yet, there is a consistent and strong effect of post-shift toluene, a remarkable consistent pattern of relative high and significant correlations with % predicted FEF25-75% (r=-0.709, p = 0.015). Post-shift m/p-xylene was also highly and significantly correlated with all % predicted spirometry parameters: FVC (r=-0.891, p = < 0.001), FEV₁ (r=-0.873, p < 0.001) and PEFR (r=-0.845, p = 0.001) and FEF25-75% r=-0.882, p = < 0.001). In multiple regression analysis when controlled for smoking, the predictors of FVC among the gas station attendants were age, height and the duration of employment as a gas station attendant (Table 4).

Exposure variables were modelled separately using predicted % of spirometry values (Table 5) and adjustment for smoking. Post-shift toluene was a negative predictor for the percentage predicted FVC in addition to the duration of employment. Exposure to benzene and m/p- and o-xylene were not significant in regression analysis and was therefore not included in the model (Table 5).

Post-shift benzene was a negative predictor for the percentage predicted FEV₁ in addition to the duration of employment (Table 5). For PEFR none of the exposure variables were included into a model.

Table 4

Regression analysis of spirometry parameters in gas station attendants (*n =* 44).
Variable	Adjusted R²	Constant	Predictors	Standardized coefficient	SE	p-value
FVC	0.651	-2.374	Age (y)	-0.027	0.006	< 0.001
			Height (cm)	0.042	0.010	< 0.001
			Duration of employment (y)	-0.017	0.007	0.020
FEV₁	0.589	-1.793	Age (y)	-0.029	0.005	< 0.001
FEV₁	0.589	-1.793	Height (cm)	0.034	0.009	0.001
PEFR	0.112	545.757	Age (y)	-2.971	1.184	0.016
FEF25-75	0.222	4.158	Age (y)	-0.038	0.010	0.001

Table 5

Regression analysis of % predicted FVC and FEV1 values in gas station attendants for BTX exposure (*n =* 20).
Variable	Adjusted R²	Constant	Predictors	Standardized coefficient	SE	p-value
% Predicted FVC	0.693	113.84	Duration of employment (y)	-0.668	0.156	0.001
% Predicted FVC	0.693	113.84	Post-shift toluene level	-0.233	0.048	< 0.001
% Predicted FEF 25–75%*	0.176	111.464	Post-shift benzene level	-0.790	0.356	0.040
* The regression equation used data from those without restriction (n = 19).

The impact of occupational exposure to BTX on lung function was evaluated by quantitative assessment of exposure to BTX as health-relevant VOCs. Our main finding was that working as a gas station attendant is consistently associated with changes in lung function with a pattern that is distinct from what was observed in office workers. In the gas station workers exposure to benzene during work hours (c.f. delta benzene) was observed to be a much more prominent risk factor, whereas in controls toluene and xylenes appear to be driving lung function changes associated with pre-shift exposure that could indicate a potential contribution from exposures outside work hours.

Study population

The gas station attendant and the control populations were comparable by age, ethnicity, anthropometric measurements including BMI and smoking status. The gas station attendants being labourers who receive a lower income compared to the non-executive office workers who were recruited as controls. The subgroups of workers for which BTX exposure data were available, were comparable (for the same parameters) to the much larger group of workers for which spirometry was successfully completed.

BTX exposure

With the collection of pre- and post-shift samples of end-exhaled air we attempted to find out to what extent BTX exposures are work-related. The rationale for this strategy is the fast kinetics of inhalation and elimination, suggesting that an increase from pre-shift to post-shift suggests exposure occurring during work hours. That this is a reasonable assumption is indicated by the strong correlations observed in gasoline station workers. The pattern of strong correlations within the BTX mixture indicates that exposure occurred during work hours, whereas the pre-shift results were clearly correlated less strongly. The high correlations in post-shift and delta results are explained by the common source of all BTX components from primarily fuel vapors and to some extent also from road vehicles where two-stroke engines of motorcycles are an important source due to incomplete combustion (IARC, 2021). The much weaker correlation with delta benzene may be explained by 5–10 times higher vapor pressure of benzene compared to toluene and xylenes. This property may affect environmental fate as well as kinetics of uptake and elimination and explain this deviation. In gas station attendants a contribution from smoking is not likely as smoking is strictly forbidden during work hours.

In our earlier report, BTX was characterised for 29 gas station attendants and 16 controls (Scheepers et al., 2019). In that study, pre- and post-shift end-exhaled air was evaluated. BTX concentrations were significantly higher among gas station attendants compared to office workers (p < 0.005). In the subgroup with spirometry values in the current analysis (20 gas station attendants and 11 controls), showed an increase in VOC levels from pre- to post-shift to be statistically significant (p < 0.03), signifying an elevated exposure to VOCs in this group of gas station attendants.

BTX can be considered both a source of indoor air pollution and outdoor air pollution ¹⁴. Toluene and xylene are commonly found in household items such as glues, paints and rubber; thus, exposure to these chemical compounds is inevitable. Benzene is primarily observed in combustion sources such as tobacco smoke and also solid fuel burning as important indoor sources. Both toluene and xylene exposure has been reported to decrease spirometry values even in residential (non-occupational) exposures. The site of toxicity is indicated to be both small and large airways with the mechanism being mediated through oxidative stress induced damage ¹⁵. That these minute exposures correlate with a reduction in lung function parameters irrespective of lack of direct and prolonged exposure to VOCs raises concern.

Gasoline station attendants and lung function

The correlation matrix indicates a strong correlation of all measured and calculated lung function parameters in controls. In gas station workers this consistent pattern of interrelationships appears to be disturbed, potentially because of different responses of lung function parameters to BTX exposures and other work-related factors. Obstructive and restrictive ventilatory patterns were similar in gas station attendants and controls who were smokers. The reduction in PEFR seen is likely to be attributable to unsatisfactory technique and effort in some attempts, and unlikely to be of any significance. The other lung function parameters were not reduced in either group, and were not significantly different between gas stations workers and controls. Measurement of airway resistance in future studies could reveal more subtle changes in lung function.

Taking together the interrelationships between BTX exposure and lung function parameters the correlation matrix in the group of gas station attendants is different from the controls. The associations observed in both gas stations attendants and controls suggest that effects on spirometry parameters are work-related. This is indicated by a consistent moderate negative correlation of % predicted FVC with post-shift and delta BTX as an indicator of the development of exposure over a shift. Associations with changes in other lung function parameters are much weaker. The more consistent negative associations with different expressions of lung function decrease with delta benzene exposure (taken as a proxy for exposure occurring during work hours) are remarkable and not observed in controls.

We used the BTX levels in end-exhaled air to correlate with the spirometry findings. Because of the high vapour pressure and poor water solubility, the retention and absorption of BTX are limited. When air concentrations are high, most of these substances have fast kinetics of uptake and elimination via the airways. This results in airways being exposed by BTX entering the lungs and a second time when these substances are eliminated via the airways as primary route of excretion. The residue leads to a build-up of BTX in the body until a steady-state is reached. When the worker is exposed to fresh air, the absorbed parent BTX (if not metabolised in the meantime) leaves the body unchanged via the lungs. This means that both during and after work, the exhaled air contains BTX, and the upper airways will be exposed during both uptake and elimination. This makes the exhaled air analysis a useful surrogate measure for airway dose (Murayama et al., 2006).

Office work and lung function

If the calculated changes from pre-shift to post-shift within each individual worker (delta) can be used as a proxy for work-related exposures it is clear that our data do not suggest BTX exposure to be associated with clinically relevant changes in lung function. A remarkable finding is the consistent and strong association of both toluene and m/p-xylene across all expressions of lung function, moderate for pre-shift and even more pronounced for post-shift. The lack of a contribution of benzene in this pattern confirms that fuels and traffic exhaust are less likely involved. Exposures to consumer products that contain toluene and xylenes may also explain these exposures and their effect on lung function, e.g., use of paint, glue and decreasing/cleaning agents with a white spirit profile of light-weight aliphatic and aromatic hydrocarbons (white-spirit type) are source of toluene and xylenes but not benzene. As we have not collected any information on the use of such products in the office environment, we cannot be sure if this exposure can be explained from work or not. As indicated by the strong negative correlation between pre-shift and post-shift values we can only suggest that is most probably derived from recent exposure, i.e., the same day.

We hypothesise that the control group was to some extent exposed to BTX outside work hours. This exposure was associated with reduced spirometry values, but to a lesser extent than the group of gas station attendants. This is reflected in somewhat lower correlations of end-exhaled air BTX levels and spirometry values. Although the BTX levels were significantly lower among the controls, even these low exposure levels were negatively correlated with the % predicted FVC. We would expect some of these correlations to become statistically significant in a larger study sample.

Employment duration

In the current study, significantly lower % predicted FVC was found among gas station attendants who were employed for 5 years or more compared to those workers with employment durations of less than 5 years. The other parameters including FEV₁/FVC ratio, % predicted FEF25-75% were lower among those employed for more than five years although not significant. Chawla and Lavania (2008) reported a significant reduction in FVC, FEV₁, PEF and FEF25-75 with increasing years of work, with FEF 25–75 being the most affected. A similar pattern has been repeatedly observed in other studies.^16–19

Context of existing knowledge

In a recently published systematic review on spirometry parameters among gas station attendants, 20 studies observed a statistically significant reduction in FVC in the exposed group compared to the controls. An additional seven studies reported lower values, although not to a statistically significant effect. The meta-analysis by the same group reported the same trend in spirometry parameters ¹⁸. A restrictive pattern in lung function among gas station attendants has been reported in previous studies ^5,19. Other studies showed an inconsistent effect on spirometry ^{16,17,20−22} with all spirometry parameters (FVC, FEV₁, FEV₁/FVC ratio, PEFR) reduced in gas station attendants compared to controls. Residents living close to fuel stations had a significantly lower FVC, and the exposure duration correlated with observed reductions in FVC values ²⁰. Similarly, in the current study, we found that duration of employment was a predictor of the FVC whereby a longer duration of employment as a gas station attendant was associated with FVC decrements but no other lung function parameters. Similar findings were reported by Rahimi et al. where persons working as petrol station attendants for more than 12 months recorded lower spirometry parameters compared to employees with less than a 12-month exposure ¹⁸. In our study, we recruited only persons exposed to fuel handling for a minimum of 12 months.

Although we did not see a statistically significant difference, a higher percentage of smokers had an abnormal spirometry pattern in both control and study groups. In a study on gas station attendants in Malta, smokers had a statistically significantly lower FVC and FEV₁ than non-smokers ²³. In our study exposure to toluene and the duration of employment were found to be negative predictors of % predicted FVC, whereas exposure to benzene was a negative predictor of % predicted FEF25-75% when controlled for smoking and all expressions of BTX exposure.

Strength and limitations

This is the first study of the Sri Lankan population where the BTX exposure levels have been correlated with spirometry values of the gas station attendants. Equally important were the exposure measurements among controls. The exposure correlation matrix of the controls is quite different from that of gas station attendants.

Important limitations in the current study are related to the small sample size due to the low response and missing values. There was random selection according to the allocation of duty on the days of exposure evaluation and the availability of acceptable spirometry recordings. Members of the research team did not select participants for either investigation based on physical or other characteristics. Only males are employed as gas station attendants in Sri Lanka, hence we did not include females also not as controls. Due to the limited availability of study participants, matching gas station attendants with controls by smoking status was not feasible. We used partial correlation and included smoking as a dependent variable in regression analysis to develop a prediction model to overcome this limitation, albeit not entirely. We recommend that our findings be reconfirmed in larger study populations to confirm associations between exposure and health outcome and allow multivariate regression analysis to better adjust for confounders. In that future work female workers should also be included if possible.

In conclusion, we did not observe changes to the lung function of any clinical relevance as a result of exposure to BTX in gas-station attendants or office workers. However, we found that obstructive responses in spirometry were most pronounced for the level of work-related benzene exposure and also associated with duration of employment. In controls a distinct pattern of associations with exposure was observed, indicating a moderate to strong association of the exposure level of toluene and m/p-xylenes. We recommend reducing long working hours and installing self-service fuel stations to minimise exposure and related adverse health effects for gas station attendants.

BTX - benzene, toluene and para, meta and ortho-xylenes

ATS - American Thoracic Society

PEFR - peak expiratory flow rate

VOC - volatile organic compounds

CPC - Ceylon Petroleum Corporation

ATD - analytical thermal desorption

FVC - Forced Vital Capacity

FEV₁- Forced Expired Volume in the first second

FEF25-75% - Forced mid expiratory flow

Ethics approval and consent to participate

Ethical approval was granted by the Ethics Review Committee of the Faculty of Medicine, University of Kelaniya, Sri Lanka. (Ref No: P/88/02/2017). All methods were carried out in accordance with relevant guidelines and regulations.Recruitment of participants followed written informed consent. The individual findings of the assessments were made available to each of the participants.

Consent for publication

Not applicable

Availability of data and materials

The datasets generated and/or analysed during the current study are not publicly available as the project is ongoing but are available from the corresponding author on reasonable request.

Competing interests

None of the authors declare competing interests.

Funding

University of Kelaniya Research grants: RP/03/04/03/01/2017 and RP/03/04/03/01/2019. The contribution of LdW was supported by the Dutch Occupational Hygiene Society (NvVA)

Authors' contributions

TW, KM, LA, PTJS, NMD were collaborators in designing, carrying out the research, data analysis and drafting the manuscript. DW, DK, DL, DDS were the physicians involved in examining the patients, data acquisition, and assisted in drafting the manuscript. JA, PR, TD, SF assisted in spirometry assessment, interpretation of the recordings and preparation of the manuscript. LDW carried out the BTX exposure assessment. DE and RW assisted in data analysis and preparation of the manuscript. All authors read and approved the final manuscript

Acknowledgement

The authors thank the gas station attendants and control their participation and their employers for facilitating the study. The authors are grateful to Maurice van Dael and Rob Anzion of the Radboudumc to prepare and support BTX exposure assessment and analysis of end-exhaled air. The authors are thankful to Jos Rooijackers and Bernadette Aalders of The Netherlands Expertise Center for Occupational and Respiratory (NECORD), Utrecht, The Netherlands for fruitful discussion of the lung function data.

All the staff members of the Department of Physiology from 2018-2020. Ms. Pavani Sandeepani for secretarial assistance.

Authors Information

TW- is a Senior Lecturer in Physiology at the Faculty of Medicine, University of Kelaniya (FOM, UOK) with a special interest in occupational exposures and how it effects the health of humans. She is a Commonwealth Medical fellow trained in autonomic Function Assessment and has won numerous awards for her research into occupational health.

KM- is a Senior Lecturer and consultant physician attached to the department of Physiology at the Faculty of Medicine, University of Kelaniya. He is in charge of the health assessment of persons recruited for studies at the FOM, UOK.

LA- is a Senior Lecturer in Physiology at the Faculty of Medicine, University of Colombo with a special interest in gastrointestinal and respiratory physiology. She is a trainer in Lung Function Testing techniques, trained at the Royal Brompton Hospital as a Commonwealth Academic Fellow. She supervises lung function testing at the Central Chest Clinic, Colombo and the Faculty of Medicine, Colombo. She is a member of the executive committee of the Physiological Society of Sri Lanka and is the outgoing Editor. Her research interests include extraoesophageal manifestations of gastroesophageal reflux disease and chronic respiratory disorders.

PTJS- is an associate professor at the Radboud University Medical Center (Radboudumc) and a registered European Registered Toxicologist (ERT) and Occupational Hygienist (RAH). His teaching is focused on human health risk assessment in both medical and science faculties at the Radboud University and in post-graduate courses on occupational toxicology and human exposure assessment in the toxicology programme. He founded the Research Lab Molecular Epidemiology at Radboudumc.

NMD- Is the Head and Chair professor in physiology at the Faculty of Medicine, University of Kelaniya, the immediate past president of the Physiological Society of Sri Lanka (PSSL) and has researched in the field of respiratory although the main interest of research has been gastroenterology.

DW- is a consultant respiratory physician and a senior lecturer attached to the faculty of Medicine, University of Kelaniya. Her main research interests include chronic respiratory diseases and obstructive sleep apnoea.

DK- is a consultant endocrinologist and a senior lecturer attached to the faculty of Medicine, University of Kelaniya. Diabetes and osteoporosis are her main research interests.

DL- is a probationary lecturer and trainee in Neurology. Her main research interests include Parkinson’s and Autonomic disorders.

DDS- is a professor in Physiology and is a consultant genetist attached to the department of Physiology at the FOM, UOK

JA, PR and TD- are Technical officers and research assistants of the department of Physiology at the FOM, UOK

SF – is a medical graduate who was a research assistant at the time of the study attached to the department of Physiology at the FOM, UOK. in spirometry assessment, interpretation of the recordings and preparation of the manuscript.

LDW- is a masters graduate from the Radboudumc who carried out the BTX exposure assessment.

DE- is the Head of the Health Data Science Unit of the FOM, UOK. His research interests include Biostatistics, Health informatics, Spatio-temporal epidemiology.

RW- is the Cadre Chair and Senior Professor and Head of the Department of Public Health, FOM, UOK. His specialization is Epidemiology and Biostatics.

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No competing interests reported.

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Association of Lung Function With Benzene, Toluene and Xylenes (Btx) in End-exhaled Air in Gas Station Attendants

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Background

Method

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Study population

End-exhaled air sampling

Analysis of end-exhaled air samples

Assessment of lung function

Statistical analysis

Results

Discussion

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