chemosensory effects during acute exposure to n-methyl-2-pyrrolidone (nmp)
TRANSCRIPT
Toxicology Letters 175 (2007) 44–56
Available online at www.sciencedirect.com
Chemosensory effects during acute exposure toN-methyl-2-pyrrolidone (NMP)
Christoph van Thriel a,∗, Meinolf Blaszkewicz a, Michael Schaper a,Stephanie A. Juran a, Stefan Kleinbeck a, Ernst Kiesswetter a,
Renate Wrbitzky b, Jurgen Stache a, Klaus Golka a, Michael Bader b
a Leibniz Research Centre for Working Environment and Human Factors at the University of Dortmund,Ardeystr. 67, D-44139 Dortmund, Germany
b Hannover Medical School, Department of Occupational Medicine, OE 5370, Carl-Neuberg-Str. 1, D-30625 Hannover, Germany
Received 8 August 2007; received in revised form 20 September 2007; accepted 21 September 2007Available online 29 September 2007
Abstract
Organic solvents are still essential in many industrial applications. To improve safety and health in the working environmentlower occupational thresholds limits have been established and less toxic substitutes were introduced. N-Methyl-2-pyrrolidone(NMP) is a versatile solvent that is used as a substitute for dichloromethane in paint strippers. Due to conflicting results, thereis a debate whether NMP causes irritations of the upper airways/eyes or not. In a human experimental study we examined thechemosensory effects of NMP under controlled conditions. Fifteen healthy males were investigated in a cross-over study. NMPvapor concentrations were 10, 40 and 80 mg/m3 for 2 × 4 h with an exposure-free lunch break of 30 min. To maximize chemosensoryeffects a peak exposure scenario (25 mg/m3 baseline, 160 mg/m3 peaks 4 × 15 min, time-weighted average: 72 mg/m3) was tested.The four different conditions were conducted with and without moderate physical workload. Chemosensory effects were measuredphysiologically by anterior rhinomanometry, eye blink rate and breathing frequency. Subjectively, ratings of acute health symptomsand intensity of olfactory and trigeminal sensations were collected repeatedly throughout the exposures. All physiological variableswere unaffected by the different NMP concentrations and even the peak exposures were non-effective on these measures. Olfactorymediated health symptoms increased dose-dependently. For these symptoms a strong adaptation was observable, especially during
the first 4 h of the exposures. Other acute symptoms were not significantly affected. Comparable to the symptoms, only olfactorysensations increased dose-dependently. Trigeminal sensations (e.g. eye and nose irritations) were evaluated as being barely detectableduring the different exposures, only during 160 mg/m3 exposure peak weak and transient eye irritation were reported. The resultsclearly suggest that NMP concentrations of up to 160 mg/m3 caused no adverse sensory irritation or undue annoyance.© 2007 Elsevier Ireland Ltd. All rights reserved.cy; Od
Keywords: Sensory irritation; Eye blink frequency; Breathing frequen∗ Corresponding author. Tel.: +49 231 1084 407;fax: +49 231 1084 308.
E-mail address: [email protected] (C. van Thriel).
0378-4274/$ – see front matter © 2007 Elsevier Ireland Ltd. All rights reservdoi:10.1016/j.toxlet.2007.09.007
or annoyance; Occupational exposure limit
1. Introduction
To advance safety and health in the work envi-
ronment the regulation of chemicals by occupationalexposure limits (OELs) is one appropriate methodof disease prevention (DFG, 2006). Additionally, leg-islative authorities may recommend the substitutioned.
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C. van Thriel et al. / Toxic
f particular chemicals by less toxic compounds. Tomprove safety and health in work environments whererganic solvents are needed for technical and indus-rial applications the German Federal Ministry of Labornd Social Affairs recommended the substitution ofichloromethane (DCM) by other compounds in a Tech-ical Rule for Hazardous Substances (TRGS 612). Inweden a comparable approach exists (Langworth et al.,001).
One frequently used substitute for DCM is N-methyl--pyrrolidone (NMP, CAS-No. 872-50-4) a cyclic amideith a weak amine-like odor. NMP is miscible with water
nd most organic solvents. In the work environmentMP has been used for surface cleaning such as graffiti
nd resin removal (Anundi et al., 1993, 2000; Bader etl., 2006; Langworth et al., 2001) or in the microelectron-cs industry (Beaulieu and Schmerber, 1991). The acuteoxicity of NMP seems to be relatively low (Payan etl., 2002) and aside from its developmental toxicity con-erns (Lee et al., 1987; Saillenfait et al., 2002, 2003) therere conflicting results concerning the irritant potency ofMP.Sensory irritation are an important and widely used
ndpoint for the regulation of chemicals (Dick andhlers, 1998; Edling and Lundberg, 2000) and it is
mportant to provide adequate data for the evaluation ofcute health effects due to irritant exposures (van Thrielt al., 2006b).
In case of NMP, Beaulieu and Schmerber (1991)nvestigated an unknown number of workers at two workites from the microelectronic industry exposed to NMPuring cleaning operations. On average air monitor-ng revealed exposure levels of 0.1–6 mg/m3 but in thelder, poorly ventilated plant airborne concentrations ofp to 276 mg/m3 were measured. The authors reportedhat “. . . via observation and by informal interview”MP levels between 62 and 70 mg/m3 were evaluatedy the workers as leading to “immediate perception,mmediately uncomfortable . . . willing to continue expo-ure only for about 30 s, minor eye irritations for thishort exposure time” (p. 878). Based of their obser-ations at these workplaces Beaulieu and Schmerberoncluded that sensory irritation, namely chronic eyerritation might be developed by workers exposed to asittle as 3 mg/m3 of NMP. Due to severe methodolog-cal problems, e.g. sample size not given, no personalir monitoring from the breathing zone, no psychome-ric scales and no statistical analysis the interpretation
f the study results is difficult. There are too many con-ounding factors and methodological shortcomings toonclude that the reported irritations can be attributedolely to NMP at this low level.etters 175 (2007) 44–56 45
Another workplace study (Langworth et al., 2001)investigating 38 graffiti removers in the Stockholm sub-way system exposed to NMP and other solvents (e.g.glycol ethers and aromatic hydrocarbons) also confirmedsome irritative effects during 8 h work shifts. Due tothe considerable co-exposures the higher prevalence ofsymptoms related to upper airway irritations amonggraffiti removers cannot be related to certain NMP con-centrations.
In contrast to workplace studies, experimental expo-sure studies can address specific effects of specificsubstances and provide useful information for the reg-ulation of chemicals by OELs. In the case of NMP theresults of a controlled 8 h exposure study with six healthymale volunteers (Akesson and Paulsson, 1997) revealedno irritative effects at concentrations of 50 mg/m3. Nei-ther pulmonary function nor nasal cavity volumes wereaffected by the three investigated concentrations of 10,25 and 50 mg/m3. Only two volunteers reported anacetone-like smell during the 50 mg/m3 condition. Theseodor perceptions were not described as being uncom-fortable. Strong odor annoyance might be unlikely atthe investigated exposure levels. Even though no irrita-tive effects were confirmed by their study, Akesson andPaulsson (1997) concluded that NMP is a mild irritantthat, at least in a concentration higher than the investi-gated 50 mg/m3, might cause sensory irritation.
The existing epidemiological data (Beaulieu andSchmerber, 1991; Langworth et al., 2001) on chemosen-sory effects of NMP is not or hardly interpretable. Incontrast, the experimental study (Akesson and Paulsson,1997) used validated and sensitive measures of sensoryirritations (Doty et al., 2004), but did not investigateconcentrations as high as under discussion for OELsetting. Recently (DFG, 2006), the Commission forthe Investigation of Health Hazards in the Work Areaof the Deutsche Forschungsgemeinschaft (DFG) hasre-established a workplace limit value (MAK-value,maximum allowable concentration in the workplace) forNMP in Germany of 20 ppm (82 mg/m3) and a corre-sponding short-term exposure limit (STEL) of 40 ppm(164 mg/m3).
The knowledge about chemosensory effects, eitherundue odor annoyance or sensory irritation due to occu-pational exposure to NMP, is far from being conclusive.To improve the database on this critical effect a compre-hensive human experimental study was carried out andexposure-scenarios comparable to workplace conditions
were investigated. These realistic scenarios includedpeak-exposures, moderate physical load, and steady-state concentrations as high as the current GermanMAK-value. A multilevel, multimethod approach was
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46 C. van Thriel et al. / Toxiused for the assessment of chemosensory effects. Toexploit the potentials of this experiment further empha-sis was put on issues of biomonitoring and urine sampleswere collected during a period of 48 h after exposureonset. The results of the biomonitoring data have beenpublished elsewhere (Bader et al., 2007a,b).
2. Materials and methods
2.1. Subjects
Sixteen healthy non-smoking male volunteers wererecruited at the University of Dortmund. These subjects wereexamined by a physician, were trained with respect to the han-dling of methods used during the study (e.g. rhinomanometer),NMP odor thresholds were measured and written informedconsent was obtained prior to the experiments. The ethicscommittee of the Leibniz Research Centre for Working Envi-ronment and Human Factors at the University of Dortmundapproved the study protocol.
After 3 test days one subject dropped out due to non-exposure related reasons. His data were excluded and allanalyses of the dependent variables are based on a sample sizeof n = 15. The mean age of the participants was 26.5 years(±2.4 years), their average height was 178 cm (±7.4 cm) andthe mean weight of the volunteers was 78 kg (±9.2 kg). Thefitness test revealed an averaged vital capacity of 4840 cm3
(±990 cm3) and the physical capacity at a heart rate of 130 bpmwas 1.84 W/kgbody weight (±0.35 W/kgbody weight). According tothese data the study population can be described as young andhealthy males with above average physical and vital capac-ity. Olfactory function was screened by means of the “Sniffin’Sticks” (Hummel et al., 1997), a standardized method forthe assessment of olfactory performance. This sensory abil-ity was assessed individually by testing of olfactory thresholdfor n-butanol, odor discrimination, and odor identification. Allsubjects performed above the criterion for hyposmia (score of30.3) of the normative sample (Kobal et al., 2000).
2.2. Exposure
The study was conducted in an exposure chamber, asecluded room built of glass and stainless steel with spatialdimensions of 4.80 m × 2.65 m × 2.27 m (≈29 m3). Inside theexposure lab four PC workplaces are located and separatedby three vertical boards. Four subjects were exposed simulta-neously. The four workplaces are equipped with 15 in. colorcomputer monitors and various response panels for symptomrating and neurobehavioral testing. During these test and scal-ing periods communication among the subjects was strictlyforbidden.
From an adjacent room a climate control unit aerated the labwith conditioned air. By means of a heater platform preciselydefined amounts of liquid NMP (provided by BASF, Germany)were vaporized and brought into the inlet airflow. The condi-tioned air was dispersed throughout the entire chamber by a
etters 175 (2007) 44–56
branched pipe system on the floor of the lab. To exhaust theNMP vapor from the lab four outlets, located at the ceiling,were used. To avoid leakage of solvent vapor into the sur-rounding laboratory the chamber was maintained at a negativepressure between 20 and 30 Pa. The average air exchange ratewas 300 m3/h. At the ceiling of the chamber four samplingdevices for online monitoring of the airborne concentrationof NMP were located and air samples were taken quasi-continuously (every 80 s). These samples were analyzed byphoto acoustic IR spectrometry (INNOVA, 1412 Photo Acous-tic Field Gas-Monitor). The results were monitored online andstored on hard disk for subsequent analyses.
2.3. Exposure scenarios
The exposure scenarios varied with respect to concentra-tion and workload. There were three time-weighted averageconcentrations (CTWA) with targeted CTWA′s of 10, 40, and80 mg/m3. While the first two concentrations were used solelywith constant time course of concentration in time (steady-state), the 80 mg/m3 was generated either as a steady-stateor as a fluctuating concentration with a basis concentra-tion of 25 mg/m3 and exposure peaks of 160 mg/m3 (Germanshort-term exposure limit, STEL). During this peak exposurescenario, potential chemosensory effects should be maximized.According to the German guidelines for the regulation of expo-sure peaks (DFG, 2006) four 160 mg/m3 peaks (duration of15 min) with intervals of 1h between successive peaks weregenerated. A 10 mg/m3 odorous control condition instead of a0 mg/m3 clean air condition was chosen to generate a conditioncapable to elicit unbiased and pure odor effects. A concentra-tion of 10 mg/m3 was selected since we obtained an averageodor threshold for NMP of approximately 8 mg/m3 in our 15participants by means of flow-olfactometry (TO7 olfactometer,ECOMA GmbH) using the method of limits according to Ger-man standards (DIN-VDI, 1999). This threshold assessmentwas conducted during the training sessions. Since no otherdata on odor thresholds for NMP was available in summaries ofodor thresholds (Devos et al., 1990; Hellman and Small, 1974;Ruth, 1986) we used this concentration as control conditionin our experiment. Fig. 1 shows the time courses of the NMPconcentration during the four different exposure scenarios.
Throughout the manuscript these exposure scenarios willbe labeled 10, 40, 80, and 25/160 mg/m3 condition.
In order to vary the uptake of NMP systematically andto simulate workplace conditions, all conditions were inves-tigated with and without additional physical workload. Theinfluence of physical stress (physical workload condition,PWL) was simulated by six 10 min periods of exercise on abicycle ergometer (75 W). To simulate mental job demandsand to substitute the ergometer exercises in the non-workload
condition, neuropsychological tests were performed. Theseconditions were called mental workload conditions (MWL).With respect to all other performed assessments of chemosen-sory effects the two workload conditions did not differ.Theresults of air monitoring are given in Table 1.
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Fig. 1. Time courses of the NMP concentration (measured by photoa-c(h
cTor(HCnrhm8rRt
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TR(l
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P
Nc
oustic spectrometry) during the four inhalational exposure scenariosaveraged across the eight sessions per CTWA condition) used in theuman exposure study.
The data given in Table 1 revealed that the targeted con-entrations were achieved with high precision and accuracy.he minimum and maximum concentration – given insteadf the standard deviation for the peak exposure scenario –evealed that the full range of the targeted concentrations25–160 mg/m3) was covered during this particular condition.owever, due to the restrictive time pattern of the peaks theTWA was about 10% below the targeted 80 mg/m3. There wereo differences between the two workload conditions of theespective exposure scenarios. During all test days the relativeumidity in the chamber was about 40% (S.D. = 5.2%) and theean temperature was around 23 ◦C (S.D. = 1.4 ◦C). During theh of each test day the temperature slightly increased in the
ange of about 2–3 ◦C due to the presence of the participants.elative humidity was not significantly affected by duration of
he exposure period.To control for order and sequence effects the sub-
ects were divided into four groups and conditions were
ermutated according to the ‘latin square’ procedure pre-enting a most concise design scheme. Four subjects eachere assigned to one of these four exposure sequences:0–40–80–25/160, 40–25/160–10–80, 80–10–25/160–40, andable 1esults of the air monitoring given as mean and standard deviation
S.D.) of the four inhalational exposure conditions and the two work-oad conditions
Target concentration
10 mg/m3 40 mg/m3 80 mg/m3 25/160 mg/m3
WL (0 W)Mean 10.58 40.86 79.85 71.82S.D. 1.22 2.16 4.27 14/170a
WL (75 W)Mean 10.37 40.37 79.65 72.30S.D. 1.20 3.96 3.58 12/175a
otes: MWL: mental workload condition; PWL: physical workloadondition.a Minima and maxima of the fluctuating concentration.
etters 175 (2007) 44–56 47
25/160–80–40–10 mg/m3. Within each sequence half of theperformed the 75 W exercise (10 min/h) bicycle ergometer ofthe PWL condition during the first session of the respective con-centration, the other half vice versa. During the subject’s nextsession this sequence was reversed. An exposure free period ofat least 1 week between two subsequent sessions was strictlyadhered to.
2.4. Ratings of chemosensory irritations
On the subjective, perceptual level the chemosensoryeffects of NMP were assessed by two different ratingapproaches. The first approach has been developed as a part ofthe Swedish Performance Evaluation System (Iregren, 1998)that has been extensively used in exposure studies investigat-ing neurobehavioral effects of various solvents (Iregren andGamberale, 1990). During the last 5 years the basic versionof the SPES subtest ‘acute symptoms’ has been extended, andsymptoms aiming at acute chemosensory health effects wereadded (Seeber et al., 2002; van Thriel et al., 2003). In principlethe procedure is as follows: one by one, 29 acute symptoms(e.g. headaches) are presented on the computer screen andtheir severity could be rated by means of a six-point categor-ical rating scale ranging from 0 (not at all) to 5 (very, verymuch). Ratings of similar items were combined into subscales.These subscales were: unspecific/pre-narcotic symptoms (foursymptoms; e.g. dizziness), olfactory symptoms (four symp-toms; e.g. nasty smell), taste symptoms (three symptoms;e.g. nasty taste), respiratory symptoms (three symptoms; e.g.shortness of breath), general irritations (three symptoms; e.g.irritation of the throat), nasal irritation (five symptoms; e.g.,itching nose), and eye irritation (seven symptoms; e.g., burningeyes).
The second rating procedure is based on the ‘labeled magni-tude scale’ (Green et al., 1996). This scale has been developedand widely used to rate the intensity of chemosensory stim-uli and the LMS mimics the ratio-like properties of magnitudeestimation scaling (Green and Shaffer, 1993), the ‘standardapproach’ for the estimation of psychophysical functions. Forthe ratings of the intensity of olfactory and trigeminal sen-sations during the exposures the following descriptors wereused: odor intensity, annoyance, eye irritation, burning, dis-gust, tickling, nasal irritation, “sneeze”, prickling, sharp, andpungent.
The SPES rating scales were presented on the computerscreen and subjects selected the respective number correspond-ing to the severity of this acute symptom on a numericalresponse panel. The LMS procedure was applied by means ofPocketPCTM (HP Jornada 540, 240 × 320 pixel) showing thescale label at the top of the screen (e.g. odor intensity), a slider
on the left side, and the six categories (ranging from: barelydetectable to strongest imaginable, numeric range: 1–1000)close to the slider. Subjects were instructed to move a smallarrow to that position of the slider that corresponds to theintensity of their current sensation.
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48 C. van Thriel et al. / ToxiBoth rating procedures were used before and after theexposures and approximately every 40 min from the onsetof the exposure at 9.00 a.m. Accordingly, for each session atotal of 14 measurements could be analyzed. By means ofthis assessment pattern ratings could be obtained at everybasis and peak concentration of the 25/160 mg/m3 condition(see Fig. 1).
2.5. Anterior active rhinomanometry (AAR)
Anterior active rhinomanometry is a method for quantita-tive measure of nasal airway resistance. Nasal congestion inresponse to an irritant should lower nasal air flow. Nasal air-flow and transnasal pressure gradient between the nostrils andthe epipharynx were measured simultaneously. In this study thecomputer-based system from Atmos Inc., Lenzkirch, Germany,was used. The flow volume is measured by ring diaphragmspiroceptor, which is adapted to the test-person by a half-mask.For anterior pressure measurement special adapters, purchasedfrom the manufacturer, were connected to the nostril accordingto nostril’s individual size and without any deformation of thenasal lobule. Transnasal pressure gradient is measured by theanterior measure method (choane to the inner pressure of themask). Flow volume and pressure gradient were calculated outof respiratory cycles recorded over 30 s by the computer-basedsystem. The thereby obtained data were checked by the so-called computer aided rhinomanometry (CAR). The left andright nostrils were measured separately and flow values attransnasal pressures of 75, 150, and 300 Pa were calculatedfrom the flow curves. AAR was measured in the morning,before exposure onset and in the evening, after the subjectsleft the chamber.
2.6. Eye blink rate
Eye blink rates were derived from electromyography(EMG) of the orbicularis oculi muscle that is responsible forthe closing of the eyelid. EMG was recorded electrophysio-logically throughout the whole sessions. To standardize visualdemands during the assessment a vigilance task (Mackworth-clock) was performed. This vigilance task lasted 25 minand was conducted four times during each test day. Thisschedule (9:15–9:40 a.m.; 11:30–11:55 a.m.; 1:45–2:10 p.m.;4:00–4:25 p.m.) was in accordance with the onset and the max-imum of the exposure peaks and thus possible effects of NMPduring exposure peaks could be investigated. For the statisticalanalyses specific sections representing the first and last 5 minof the four respective 25-min vigilance tests were selected.
2.7. Breathing rate
The breathing frequency was measured by means of respi-ratory inductive plethysmography (RIP). A flexible breast beltwas put around the subject’s chest. Within this flexible belt aforked wire is used as a coil. The movement of the chest changesthe inductance of this “coil” and the biosignal recorder converts
etters 175 (2007) 44–56
these changes into electronic signals that were digitized andstored on CF-cards. Based on the recorded oscillating breath-ing curve the mean individual breathing rate was calculated.To minimize artefacts from body movements, the analyses ofthe breathing rate was limited to the time intervals also usedfor the analyses of the eye blink rate.
2.8. Statistics
Since the symptom ratings were ordinal-scaled with poten-tial values from 0 to 5, non-parametric tests were used.Separate Page’s trend test (Page, 1963) were used to iden-tify differences between the four exposure scenarios (10, 40,80, 25/160 mg/m3) either during PWL or MWL conditions.This non-parametric trend test allows for multiple compar-isons between ordered correlated variables. Based on therespective time-weighted average exposures of the investi-gated exposure conditions, we tested for the following trend:10 < 40 < 25/160 < 80 mg/m3. If the results of Page’s trend testsconfirmed significant effects of the ascending concentrationsteps, pairwise comparisons were calculated to identify sig-nificant differences between the exposure scenarios. Finally,the different workload conditions were compared by Wilcoxontests testing each exposure scenario separately (e.g. 10 mg/m3
PWL vs. 10 mg/m3 MWL).The LMS data, coming from continuous scales, was ana-
lyzed in a two-stage approach. In a first step Wilcoxon testswere use to compare the pre-exposure ratings with medianratings given during the different exposure conditions. Dueto skewed distributions, especially of the pre-exposure rat-ings this non-parametric test was used. Those intensity ratingsthat were not significantly elevated during any exposure wereexcluded from the second step of the analysis. In that sub-sequent step dose-response effects were analyzed by meansof repeated measurements analysis of variance (ANOVA). A2 × 4 × 2 × 6-factorial model was applied to the twelve rat-ings given during the exposures. In detail, these factors were:exposure condition (10, 40, 80, 25/160 mg/m3), time of day(morning vs. afternoon), measuring time (six times per half-shift) and workload (mental vs. physical). In order to analyzethe time courses of the ratings in-depth the three steady-stateconditions and the peak exposure scenario were analyzed sepa-rately. Planned comparisons were used to describe the impact ofexposure peaks and a reduced model (2 × 3 × 2 × 6-factorial)was applied to the data of the three constant exposure condi-tions.
The rhinomanometry, eye blink rate and breathing fre-quency were analyzed by means of repeated measurementsanalysis of variance (ANOVA) using a 4 × 4 × 2 × 2-factorialmodel. These factors were: exposure condition (10, 40, 80,25/160 mg/m3), measuring time (four times), section (begin-
ning vs. end of vigilance task) and workload (mental vs.physical). The factor section also represents the impact of expo-sure peaks during the 25/160 mg/m3 condition. Post hoc testsof this factor were used to describe the impact of exposurepeaks.
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A significance level of α = 0.05 was chosen to test the nullypothesis of no differences between population means.
. Results
After the exposure session’s subjects spontaneouslyescribed the quality of the NMP odor as weak, some-hat sweet, like dry-cleaning and to some extent fishy.he standardized assessments yielded the following
esults.
.1. Acute symptoms (SPES)
In general almost no acute health symptoms wereeported during the eight exposures. Six of the sevenymptom subscales yielded average ratings of 0 (not atll) or 1 (hardly at all) throughout all investigated expo-ure and workload conditions. Page’s trend tests revealedo significant differences for ratings of unspecific/ pre-arcotic, taste, respiratory symptoms, general irritations,asal irritation or eye irritation. Within the four CTWAteps these symptom subscales showed no significantifferences between physical and mental workload con-itions.
Only for olfactory symptoms Page’s trend testsndicated significant differences between the ratingsiven during the four CTWA steps for exposures withental (L = 412, p < 0.001) and physical workload
L = 395, p < 0.05). While for the mental workloadonditions this trend was also confirmed by signifi-
ant pairwise comparisons, the pairwise comparisonsf the physical workload condition yielded insignif-cant differences. Therefore, Fig. 2 only shows theroportion of participants reporting different severityig. 2. Stacked bar chart of the olfactory symptoms (sensation of badir (quality), nasty smell, sensation of unpleasent smell, stink) givenuring the different exposure conditions with mental workload.
etters 175 (2007) 44–56 49
categories (not at all, hardly at all, somewhat, rathermuch) during the four concentration steps with mentalworkload.
The simple stacked bar chart shows a concentra-tion dependent increase of higher severity categoriesfor the three steady-state conditions. The significanceof this dose-dependency was confirmed by the twoWilcoxon tests comparing the 10 and 40 mg/m3 con-dition (z = −2.12, p = 0.03) and the 40 and 80 mg/m3
condition (z = −2.65, p < 0.01). While the average rat-ings given during the 25/160 mg/m3 peak condition werealso significant higher than during the 10 mg/m3 odor-ous control condition (z = −2.76, p < 0.01), they were notsignificantly different from either the 40 or 80 mg/m3
conditions. Like for the six other symptom subscales,the final comparisons of the two workload alternativeswithin the four CTWA steps revealed no significant dif-ferences for ratings of acute olfactory mediated healthsymptoms.
3.2. Ratings of chemosensory sensations
The profiles of chemosensory sensations elicited byNMP during the eight different exposures are given inFig. 3.
It is obvious from Fig. 3 that under the investigatedconditions only very weak intensities of chemosen-sory sensations were reported. Even during conditionsincluding exposure peaks of 160 mg/m3 of NMP sev-eral, especially trigeminal mediated sensations werebarely detectable. The presence of physical or mentalworkload altered the profiles only slightly. The odor ofNMP seemed to be detectable during all conditions andtogether with annoyance this sensation increased almostdose-dependent. Such a general dose-dependency wasnot observed for nose and eye irritations. While ratingsof eye irritation did not clearly differ between the 40,80 and 25/160 mg/m3 condition, nose irritation showedsome dose-dependency during MWL but not for PWLconditions, where nose irritation were highest during to40 mg/m3 condition. However, except for odor intensityand annoyance the ratings are very low and subsequentlyit will be analyzed whether these ratings are different forbaseline level obtained prior to the exposures.
The non-parametric analyses showed that during noneof the eight investigated exposure scenarios the fivesensations nauseous, prickling, burning, “sneeze”, andtickling, were significantly different from the ratings
obtained at baseline. The results of the Wilcoxon testsfor the remaining items are given in Table 2.During the 10 mg/m3 conditions the six remainingsensations were not consistently elevated. Some incon-
50 C. van Thriel et al. / Toxicology Letters 175 (2007) 44–56
the fou
Fig. 3. Profile of the intensity of chemosensory sensations elicit bypanel) and physical workload (lower panel).sistencies between the two workload conditions wereprobably caused by slightly lower baseline ratings (e.g.1 vs. 14 for odor intensity). As visible from Fig. 3,the trigeminal mediated sensations pungent and sharpwere only slightly different from one and thus significantdifferences from baseline occurred only sporadically.The intensity of nose and eye irritations differed frombaseline only infrequently. Especially during the highest
steady-state exposure no consistent increase of these sen-sations could be confirmed. Somewhat stronger effects,with z-values around 3 were only observable for the twosensations odor intensity and annoyance. The signifi-r different NMP exposure for sessions with mental workload (upper
cant elevation of these ratings was consistent betweenthe workload conditions. According to these results thetwo-step ANOVAs were only calculated for rating ofodor intensity and annoyance. Furthermore, the intensityof nose and eye irritations were used separately in orderto analyze the impact of exposure peaks on trigeminalsensations.
The overall ANOVA – including the 25/160 mg/m3
condition – revealed that odor intensity differed signifi-cantly in relation to exposure condition (F(3,42) = 16.98,p < 0.001) and measuring time (F(5,70) = 13.90,p < 0.001). Workload and time of day did not sig-
C. van Thriel et al. / Toxicology Letters 175 (2007) 44–56 51
Table 2Results of the Wilcoxon test comparing pre-exposure ratings with those obtained during the eight inhalational exposure conditions
10 mg/m3 40 mg/m3 80 mg/m3 25/160 mg/m3
MWL PWL MWL PWL MWL PWL MWL PWL
Pungentz-Value −2.20 −2.37 −2.37 −2.37 −2.37p 0.03 0.02 0.02 0.02 0.02
Sharpz-Value −2.37 −2.20 −2.20 −2.20p 0.02 0.03 0.03 0.03
Nose irritationsz-Value −2.52 −2.52 −2.80 −2.80p 0.01 0.01 0.01 0.01
Eye irritationz-Value −2.37 −2.52 −2.52 −2.80 −2.13p 0.02 0.01 0.01 0.01 0.03
Odor intensityz-Value −2.55 −3.11 −2.95 −3.11 −3.41 −3.12 −3.41p 0.01 0.00 0.00 0.00 0.00 0.00 0.00
Annoyancez-Value −2.67 −2.35 −2.67 −3.30 −2.67 −2.54 −2.79
0.01
N
ndaicatt
Frtta
p 0.01 0.02
otes: MWL: mental workload, PWL: physical workload.
ificantly affect the ratings and therefore descriptiveata was collapsed across these factors. Fig. 4 showsn almost linear increase of odor intensity withncreasing NMP concentration for the steady-state
onditions. Assuming that 25/160 mg/m3 condition hadCTWA of 72 mg/m3 the average rating given duringhe peak exposure scenario would fit into a linearrend.
ig. 4. (Left side) Means and standard deviations of odor intensityated during the four different exposure conditions; (right side) varia-ion of means and standard deviations of odor intensity ratings acrosshe six measuring times per half-shifts, collapsed across morning andfternoon and averaged across exposure conditions.
0.00 0.01 0.01 0.01
It is obvious for Fig. 4 (right side) that the sig-nificant main effect of the factor measuring time wascaused by the exposure peaks located at the 2nd and5th measurement of each half-shift (see Fig. 1). Accord-ing to this resounding impact of peak exposure on thetime courses of chemosensory sensations any signif-icant interactions of the factors were not interpretedyet, but in the second step of the analysis of vari-ance. The second ANOVA confirmed the main effectsof CTWA (F(2,28) = 18.84, p < 0.001) and measuringtime (F(5,70) = 4.90, p < 0.01) as well as their interac-tion (F(10,140) = 2.61, p = 0.01). Moreover, the analysisrevealed a significant triple interaction on CTWA, time ofday and measuring time (F(10,240) = 2.00, p = 0.04). Thedifferentiated time courses of the three steady-state con-ditions either in the morning or in the afternoon are givenin Fig. 5.
Fig. 5 shows that across the 4 h the first half-shift(morning) odor intensity decreased for the 40 and80 mg/m3 condition and the decrease was stronger atthe higher CTWA condition. In the afternoon the CTWA-dependent differences in the perception of the NMP odorremained but additional effects of the exposure duration
did not occur.The second ANOVA analyzed annoyance ratingsand effects comparable to those previously describedfor odor intensity occurred. The exposure conditions
52 C. van Thriel et al. / Toxicology L
Fig. 5. Time courses of odor intensity across the two 4 h half-shifts ofthe three steady-state exposure conditions.
clearly affected annoyance (F(3,42) = 8.59, p < 0.001),with higher annoyance ratings at higher CTWA levels.Like odor intensity, the annoyance rating peaked atthe 2nd and 5th measurement of the half-shifts andthus the factor measuring time had a significant effect(F(5,70) = 6.87, p < 0.001). Again, workload and time ofday did not significantly affect the ratings. In contrast toodor intensity the steady-state ANOVA revealed no dif-ferentiated time course of the annoyance rating and onlythe main effect of CTWA was confirmed (F(2,28) = 10.30,p < 0.001).
The mean ratings of odor intensity, annoyance, noseand eye irritations given during the different measur-
ing times of the 25/160 mg/m3 exposures are given inFig. 6.While odor intensity and annoyance clearly mim-icked the time course of the NMP concentrations (see
Fig. 6. Time courses of the mean intensity ratings odor intensity,annoyance, eye irritations and nose irritations across the two 4 hhalf-shifts given during the 25/160 mg/m3 peak exposure conditions(collapsed across mental and physical workload conditions).
etters 175 (2007) 44–56
Fig. 1) this pattern was less distinct for nose andeye irritations. The four planned comparisons of rat-ings given at peak concentrations (9.40 a.m., 11.55 a.m.,2.10 p.m., 4.25 p.m.) with neighboring rating given atthe basis concentration of 25 mg/m3 were highly sig-nificant for both olfactory sensations. For eye irritationonly the first peak showed a significant effect on theratings (F(1,14) = 8.62, p = 0.01). Nasal irritation weresignificantly elevated during the second (F(1,14) = 6.05,p = 0.03), third (F(1,14) = 4.79, p = 0.05) and fourth expo-sure peak (F(1,14) = 8.54, p = 0.01).
3.3. Rhinomanometry, eye blink rate, and breathingfrequency
The nasal flow of the participants, as measure byrhinomanometry, was significantly lower after the expo-sures (F = 6.653, p = 0.02). The ANOVA revealed thatthis decreasing flow was neither affected by the CTWAcondition, nor by the different types of workload. Therespective flow changes of the four exposure condi-tions were: �Flow = −42 ml/s at 10 mg/m3, −37 ml/sat 40 mg/m3, −36 ml/s at 80 mg/m3, and −10 ml/s at25/160 mg/m3.
The eye blink rates were not affected by the fac-tor exposure conditions. Exposure peaks did not affectthe blinking rates. The ANOVAs revealed only onesignificant main effect of the four repetitions of this (psy-cho)physiological measurement (F = 3.04, p = 0.04). Onaverage, the mean eye blink rates increase from16.0 min−1 (1st measurement), to 17.2 min−1 (2nd), to17.3 min−1 (3rd), and finally 17.7 min−1 (4th) at the endof the test day.
The ANOVA of the breathing frequency revealedthree significant main effects of (a) the two workloadconditions (F = 5.14, p = 0.04), (b) the four repetitionsof this (psycho)physiological measurement within a ses-sion (F = 13.51, p < 0.01), and (c) the two sections at thefirst and last 5 min of each 25-min assessment period(F = 24.86, p < 0.01). There was no effect of the CTWAcondition, either as a main effect or in interaction withany of the other factors used in this statistical model. Themean breathing rates together with their 0.95 confidenceinterval, subject to every significant factor, are given inFig. 7.
On average, the breathing rates were slightly higherduring the measurements obtained in the exposuresessions with 75 W physical workload. Across a test
day, the breathing rates of the subjects increasedfrom about 15.5 to 16.7 breaths per minute. Withinthe 25-min periods of the measurements the breath-ing rates decreased. Although these effects were
C. van Thriel et al. / Toxicology L
Fig. 7. Means and 95%-confidence intervals of the breathing fre-quency observed for the different workload conditions, across the fourset
sii
4
saNtwttcEnwttwesdpigpsm
tandardized measuring times (25-min vigilance task) during the 8 hxposure period, and for the first and last 5 min (analyzed sections) ofhese four standardized measuring times.
tatistically significant the sizes of the changes of breath-ng rates were relatively small and physiologicallyrrelevant.
. Discussion
Summing up the results of this experimental expo-ure study we have shown that in concentrations as highs current European OELs and STELs (DFG, 2006)MP caused predominantly odor effects. The magni-
udes of these olfactory mediated effects were relativelyeak and during simulated work shifts adaptation to
he odor of NMP occurred. The intensity ratings ofrigeminally mediated sensations were not systemati-ally elevated during the four different NMP exposures.specially burning sensations mediated by unmyeli-ated C-fibers of the trigeminal nerve (Hummel, 2000)ere not elicited by acute NMP exposures. In contrast,
he two sensations pungent and sharp, physiologicallyransmitted by myelinated Adelta-fibers (Hummel, 2000),ere at least occasionally different from baseline (pre-
xposure ratings). Nevertheless, none of these subjectiveigns of sensory irritations were elicit concentration-ependent. In line with those perceptional results, thehysiological measures assessing ocular and intranasalrritations were not significantly affected by the investi-
ated NMP concentrations. Moreover, even if exposureeaks of 160 mg/m3 were used during the varying expo-ure scenarios the blinking frequency, a very sensitivearker (Kiesswetter et al., 2005) of exposure peaks, wasetters 175 (2007) 44–56 53
not elevated. Accordingly, the results of the Akessonand Paulsson study (1997), where no sensory irritationwere revealed during acute experimental exposures to50 mg/m3 of NMP could be replicated and expanded toeven higher concentrations. Therefore, the results of thepresent study provide first evidence that acute NMP con-centrations up to 160 mg/m3 do not cause severe sensoryirritation.
In contrast, the strong irritation suggested by theBeaulieu and Schmerber study (1991) could not be con-firmed by the results of the present study and the Akessonand Paulsson data (1997). Confounding in workplacestudies can be controlled in experimental exposure stud-ies investigating healthy human volunteers. However,some concerns exist about the responses of naıve volun-teers in comparison to occupationally exposed workersthat might habituate to the odor and mild irritation duringthere working life (Ihrig et al., 2006). Therefore, exper-imental exposure studies with naıve volunteers mightoverestimate the relevant trigeminal potency of a chem-ical used in the workplace. Regarding this problem, ithad been shown that workers occupationally exposedto ammonia reported less irritations than naıve volun-teers (Ihrig et al., 2006). Several psychophysical studiesshowing higher odor and trigeminal detection thresholdsin workers than controls, solely for the occupationallyused substance (Dalton et al., 2003; Wysocki et al.,2003), lead to the suggestion that adaptation might beresponsible for the reduction of the symptom reports.Regardless of the underlying mechanisms, the describeddifference between workers and naıve volunteers mightalso be relevant for the interpretation of our results. Theweak olfactory effects that we reported in our experi-mental exposure study might be an overestimation ofthe chemosensory effects of acute NMP exposures inworkers increasing the margin of safety for disease pre-vention.
The valid assessment of sensory irritations and espe-cially the differentiation from pure odor effects iscrucial (Arts et al., 2006; Dalton, 1996, 2001; vanThriel et al., 2006b). Simple self-reports, as used byBeaulieu and Schmerber (1991), might be subjected tosevere bias due to non-sensory factors (Dalton, 2002;Smeets and Dalton, 2005). Such factors are beliefsabout health risks due to chemicals (Dalton, 2002),personality traits related to chemical exposures, likeself-reported pollutant reactivity (Shusterman, 2002),or experiences with the local irritant as discussed in
the previous section. Experience-based effects are notrestricted to occupational exposures. Pleasantness rat-ings were affected by the previous experience of amalodor in a controlled chamber with two successive
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54 C. van Thriel et al. / Toxisessions (Knasko, 1992). Additionally, the selectionof adequate and validated rating scales is crucial forthe interpretation of subjective data on chemosensoryeffects.
4.1. Validity of the measurements
In previous studies we could show that, compared tosymptom ratings, the LMS rating procedure we used inthe present study is less susceptible to influences frompersonality factors like self-reported chemical sensitiv-ity (van Thriel et al., 2005). One crucial proof of thevalidity of a scaling procedure is the existence of a suffi-cient concentration–effect relationship. Psychophysicalfunctions according to Stevens’ power law (Ψ = k × Φ�),where Ψ is the perceived intensity (e.g. odor intensity)and Φ is the concentration of the investigated localirritant, can be used to test concentration–effect rela-tionships in scaling experiment. Such an experiment,investigating six local irritants, had been conducted forthe LMS scaling procedure used in the present study (vanThriel et al., 2006a). The LMS could be predicted by theapplied concentrations with high accuracy as reflectedby R2 values of the psychophysical functions rangingfrom 0.86 to 0.99 (van Thriel et al., 2006a). Furthermore,another experimental exposure study, investigating acuteirritations due to iso-propanol exposures, used similarscaling procedures and a good concentration–effect rela-tionships were confirmed (Smeets et al., 2002). Thevalidation of the ordinal ratings from SPES questionnairedata was also confirmed in an experimental exposurestudy. Again the concentration–effect relationship wasused for validity check. The sinusoidal time course ofthe concentrations of several chemical, all investigatedin 4 h exposure experiments, was used to predict olfac-tory symptoms, nasal and eye irritations (van Thriel etal., 2003). The non-linear regression revealed R2 valuesof 0.86 for olfactory symptom, 0.91 for nasal irrita-tion and 0.51 for eye irritation. These goodness-of-fitparameters suggest that if a substance is really irritat-ing the SPES symptom questionnaire is a valid toolto screen for sensory irritation. This proof of validityfor the subjective scaling methods is neither availablefor the Akesson and Paulsson study (1997) nor for theBeaulieu and Schmerber study (1991). Therefore, theresults of the rating methods of our study can be con-sidered as more valid and quality ensured than thoseof the other studies addressing chemosensory effects
of NMP.The physiological measures were in line with theexpert recommendation for the assessment of healtheffects of local irritants (Doty et al., 2004). Especially
etters 175 (2007) 44–56
the analysis of eye blink rate has been widely used toobjectify trigeminal stimulations unbiased from olfac-tion (Emmen et al., 2003; Iregren et al., 1993; Kleno andWolkoff, 2004; Walker et al., 2001). The EMG techniquethat we used showed good validity in an experimen-tal exposure experiment investigating 2-ethylhexanol(Kiesswetter et al., 2005). Both, general concentra-tion dependency of the three investigated time-weightedaverage concentrations as well as strong increases ofthe eye blink rate during exposure peaks could beconfirmed. By rhinomanometry nasal congestion afterexposures to chlorine could be confirmed (Shusterman etal., 2003), and therefore, also this method can be consid-ered as sensitive to identify intranasal effects of trigemialstimulation. In conclusion, the validity of our methodsappeared to be sufficient and the obvious discrepancy ofour results to those of the Beaulieu and Schmerber study(1991) cannot be explained by the usage of insensitivemethods.
4.2. Adaptation and other effects on breathing
Olfactory sensations are subjected not only to long-term adaptation (Dalton and Wysocki, 1996), theolfactory system is designed to adapt quickly to rel-atively short stimulation periods (Dalton, 2000). Theconcentration-dependent adaptation that we reported forthe odor intensity ratings confirmed that mainly olfactorypathways contributed to this effect. For the evaluation ofsubjective symptoms in studies aiming at chemosensoryeffects the analysis of time courses has been recom-mended (van Thriel et al., 2005) to evaluate the adversityof the observed effect. In contrast to olfactory mediatedsensations, reported irritation are subjected to temporalintegration (Wise et al., 2004, 2005). Accordingly, dur-ing constant stimulation, as given during our steady-stateexposures, the absence of adaptation in subjective ratingswould be a hint of trigeminal co-stimulation. The tem-poral analysis of our data provides additional support forthe conclusion that NMP caused only olfactory-mediatedeffects.
The analysis of the breathing rates revealed someeffects unrelated to the exposure scenarios. Even thoughthe magnitudes of these effects were really small, theyappear to be physiologically sound and can be interpretedas proof of validity. Residual effects of the physicaleffort during ergometer exercise might be responsiblefor the slight increase of the breathing rate during PWL
conditions. This interpretation can be supported by thedecrease of the breathing rate from the beginning to theend of the analyzed time segment. During this periodsubjects performed sedentary work and general relax-
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tion might occur. We observed similar effects duringnother 4 h exposure experiment (Haumann et al., 2003).he increase of the breathing rate from the beginning to
he end of the 8 h exposure session might be related tonspecific changes in climate conditions, most likely thelight increase of room temperature. However, due to their exchange rate of 10 times per hour increasing levelsf CO2 can be ruled out being causative for this effect.aken together these results showed the applied physio-
ogical measures were sensitive to relevant changes andhus capable to identify irritations of the upper respira-ory tract.
. Conclusion
Due to the consistency of our data with the resultsf a previous environmental chamber study (Akessonnd Paulsson, 1997), the higher degree of controlling foro-variables, the usage of physiological measures (e.g.hinomanometry) for the assessment of sensory irrita-ions and the proven validity of the rating techniques, theresent study allowed the conclusion that NMP causedo adverse sensory irritation in concentrations coveredy current OEL values (DFG, 2006).
cknowledgement
This study was financially supported by grants ofhe NMP Producers Group, c/o Bergeson & Campbell,
ashington, DC, USA.
eferences
kesson, B., Paulsson, K., 1997. Experimental exposure of malevolunteers to N-methyl-2-pyrrolidone (NMP): acute effects andpharmacokinetics of NMP in plasma and urine. Occup. Environ.Med. 54, 236–240.
nundi, H., Lind, M.L., Friis, L., Itkes, N., Langworth, S., Edling, C.,1993. High exposures to organic solvents among graffiti removers.Int. Arch. Occup. Environ. Health 65, 247–251.
nundi, H., Langworth, S., Johanson, G., Lind, M.L., Akesson, B.,Friis, L., Itkes, N., Soderman, E., Jonsson, B.A., Edling, C., 2000.Air and biological monitoring of solvent exposure during graffitiremoval. Int. Arch. Occup. Environ. Health 73, 561–569.
rts, J.H., de Heer, C., Woutersen, R.A., 2006. Local effects in therespiratory tract: relevance of subjectively measured irritation forsetting occupational exposure limits. Int. Arch. Occup. Environ.Health, 1–16.
ader, M., Rosenberger, W., Rebe, T., Keener, S.A., Brock, T.H.,Hemmerling, H.J., Wrbitzky, R., 2006. Ambient monitoring and
biomonitoring of workers exposed to N-methyl-2-pyrrolidone inan industrial facility. Int. Arch. Occup. Environ. Health 79, 357–364.ader, M., Wrbitzky, R., Blaszkewicz, M., Schaper, M., van Thriel,C., 2007a. Human volunteer study on the inhalational and der-
etters 175 (2007) 44–56 55
mal absorption of N-methyl-2-pyrrolidone (NMP) from the vapourphase. Arch. Toxicol., doi:10.1007/s00204-007-0230-5.
Bader, M., Wrbitzky, R., Blaszkewicz, M., van Thriel, C., 2007b.Human experimental exposure study on the uptake and urinaryelimination of N-methyl-2-pyrrolidone (NMP) during simulatedworkplace conditions. Arch. Toxicol. 81, 335–346.
Beaulieu, H.J., Schmerber, K.R., 1991. M-Pyrol(TM) (NMP) use inthe microelectronics industry. Appl. Occup. Environ. Hyg. 6, 874–880.
Dalton, P., 1996. Odor perception and beliefs about risk. Chem. Senses21, 447–458.
Dalton, P., 2000. Psychophysical and behavioral characteristics ofolfactory adaptation. Chem. Senses 25, 487–492.
Dalton, P., 2001. Evaluating the human response to sensory irritation:implications for setting occupational exposure limits. AIHAJ 62,723–729.
Dalton, P., 2002. Odor, irritation and perception of health risk. Int.Arch. Occup. Environ. Health 75, 283–290.
Dalton, P., Wysocki, C.J., 1996. The nature and duration of adapta-tion following long-term odor exposure. Percept. Psychophys. 58,781–792.
Dalton, P., Cowart, B., Dilks, D., Gould, M., Lees, P.S., Stefaniak, A.,Emmett, E., 2003. Olfactory function in workers exposed to styrenein the reinforced-plastics industry. Am. J. Ind. Med. 44, 1–11.
Devos, M., Patte, F., Rouault, J., Laffort, P., Van Gemert, L.J., 1990.Standarsized Human Olfactory Thresholds. IRL Press, Oxford.
DFG, 2006. List of MAK and BAT Values 2006. Wiley-VCH VerlagGmbH, Weinheim.
Dick, R.B., Ahlers, H., 1998. Chemicals in the workplace: incorpo-rating human neurobehavioral testing into the regulatory process.Am. J. Ind. Med. 33, 439–453.
DIN-VDI, 1999. Messen und Bewerten von Geruchen: TechnischeRegeln. Beuth, Berlin.
Doty, R.L., Cometto-Muniz, J.E., Jalowayski, A.A., Dalton, P., Kendal-Reed, M., Hodgson, M., 2004. Assessment of upper respiratorytract and ocular irritative effects of volatile chemicals in humans.Crit. Rev. Toxicol. 34, 85–142.
Edling, C., Lundberg, P., 2000. The significance of neurobehavioraltests for occupational exposure limits: an example from Sweden.Neurotoxicology 21, 653–658.
Emmen, H.H., Muijser, H., Arts, J.H., Prinsen, M.K., 2003. Humanvolunteer study with PGME: eye irritation during vapour exposure.Toxicol. Lett. 140/141, 249–259.
Green, B.G., Shaffer, G.S., 1993. The sensory response to capsaicinduring repeated topical exposures: differential effects on sensationsof itching and pungency. Pain 53, 323–334.
Green, B.G., Dalton, P., Cowart, B., Shaffer, G., Rankin, K., Higgins,J., 1996. Evaluating the ’Labeled Magnitude Scale’ for measuringsensations of taste and smell. Chem. Senses 21, 323–334.
Haumann, K., Kiesswetter, E., van Thriel, C., Blaszkewicz, M., Golka,K., Seeber, A., 2003. Breathing and heart rate during experimen-tal solvent exposure of young adults with self-reported multiplechemical sensitivity (sMCS). Neurotoxicology 24, 179–186.
Hellman, T.M., Small, F.H., 1974. Characterization of the odor prop-erties of 101 petrochemicals using sensory methods. J. Air Pollut.Control Assoc. 24, 979–982.
Hummel, T., 2000. Assessment of intranasal trigeminal function. Int.
J. Psychophysiol. 36, 147–155.Hummel, T., Sekinger, B., Wolf, S.R., Pauli, E., Kobal, G., 1997. ’Snif-fin sticks’: olfactory performance assessed by the combined testingof odor identification, odor discrimination and olfactory threshold.Chem. Senses 22, 39–52.
cology L
56 C. van Thriel et al. / ToxiIhrig, A., Hoffmann, J., Triebig, G., 2006. Examination of the influenceof personal traits and habituation on the reporting of complaints atexperimental exposure to ammonia. Int. Arch. Occup. Environ.Health 79, 332–338.
Iregren, A., 1998. Computer-assisted testing. In: Costa, L.G., Manzo,L. (Eds.), Occupational Neurotoxicology. CRC Press LLC, BocaRaton, pp. 213–231.
Iregren, A., Gamberale, F., 1990. Human behavioral toxicology. Cen-tral nervous effects of low-dose exposure to neurotoxic substancesin the work environment. Scand J. Work Environ. Health 16 (Suppl.1), 17–25.
Iregren, A., Lof, A., Toomingas, A., Wang, Z., 1993. Irritation effectsfrom experimental exposure to n-butyl acetate. Am. J. Ind. Med.24, 727–742.
Kiesswetter, E., van Thriel, C., Schaper, M., Blaszkewicz, M., See-ber, A., 2005. Eye blinks as indicator for sensory irritation duringconstant and peak exposures to 2-ethylhexanol. Environ. Toxicol.Pharmacol. 19, 531–541.
Kleno, J., Wolkoff, P., 2004. Changes in eye blink frequency as ameasure of trigeminal stimulation by exposure to limonene oxi-dation products, isoprene oxidation products and nitrate radicals.Int. Arch. Occup. Environ. Health 77, 235–243.
Knasko, S.C., 1992. Ambient odor’s effect on creativity, mood, andperceived health. Chem. Senses 17, 27–35.
Kobal, G., Klimek, L., Wolfensberger, M., Gudziol, H., Temmel, A.,Owen, C.M., Seeber, H., Pauli, E., Hummel, T., 2000. Multicenterinvestigation of 1,036 subjects using a standardized method for theassessment of olfactory function combining tests of odor identi-fication, odor discrimination, and olfactory thresholds. Eur. Arch.Otorhinolaryngol. 257, 205–211.
Langworth, S., Anundi, H., Friis, L., Johanson, G., Lind, M.L., Soder-man, E., Akesson, B.A., 2001. Acute health effects common duringgraffiti removal. Int. Arch. Occup. Environ. Health 74, 213–218.
Lee, K.P., Chromey, N.C., Culik, R., Barnes, J.R., Schneider, P.W.,1987. Toxicity of N-methyl-2-pyrrolidone (NMP): teratogenic,subchronic, and two-year inhalation studies. Fundam. Appl. Toxi-col. 9, 222–235.
Page, E.B., 1963. Ordered hypotheses for multiple treatments—a sig-nificance test for linear ranks. J. Am. Stat. Assoc. 58, 216–230.
Payan, J.P., Beydon, D., Fabry, J.P., Boudry, I., Cossec, B.,Ferrari, E., 2002. Toxicokinetics and metabolism of N-[14C]methylpyrrolidone in male Sprague–Dawley rats. A saturableNMP elimination process. Drug Metab. Dispos. 30, 1418–1424.
Ruth, J.H., 1986. Odor thresholds and irritation levels of several chem-
ical substances: a review. Am. Ind. Hyg. Assoc. J. 47, A142–A151.Saillenfait, A.M., Gallissot, F., Langonne, I., Sabate, J.P., 2002. Devel-opmental toxicity of N-methyl-2-pyrrolidone administered orallyto rats. Food Chem. Toxicol. 40, 1705–1712.
etters 175 (2007) 44–56
Saillenfait, A.M., Gallissot, F., Morel, G., 2003. Developmentaltoxicity of N-methyl-2-pyrrolidone in rats following inhalationexposure. Food Chem. Toxicol. 41, 583–588.
Seeber, A., van Thriel, C., Haumann, K., Kiesswetter, E., Blaszkewicz,M., Golka, K., 2002. Psychological reactions related to chemosen-sory irritation. Int. Arch. Occup. Environ. Health 75, 314–325.
Shusterman, D., 2002. Individual factors in nasal chemesthesis. Chem.Senses 27, 551–564.
Shusterman, D., Balmes, J., Avila, P.C., Murphy, M.A., Matovinovic,E., 2003. Chlorine inhalation produces nasal congestion in aller-gic rhinitics without mast cell degranulation. Eur. Respir. J. 21,652–657.
Smeets, M., Dalton, P., 2005. Evaluating the human response to chem-icals: odor, irritation and non-sensory factors. Environ. Toxicol.Pharmacol. 19, 581–588.
Smeets, M.A., Maute, C., Dalton, P.H., 2002. Acute sensory irritationfrom exposure to isopropanol (2-propanol) at TLV in workers andcontrols: objective versus subjective effects. Ann. Occup. Hyg. 46,359–373.
van Thriel, C., Seeber, A., Kiesswetter, E., Blaszkewicz, M., Golka, K.,Wiesmuller, G.A., Physiological, 2003. psychological approachesto chemosensory effects of solvents. Toxicol. Lett. 140/141,261–271.
van Thriel, C., Kiesswetter, E., Schaper, M., Blaszkewicz, M., Golka,K., Seeber, A., 2005. An integrative approach considering acutesymptoms and intensity ratings of chemosensory sensations dur-ing experimental exposures. Environ. Toxicol. Pharmacol. 19,589–598.
van Thriel, C., Schaper, M., Kiesswetter, E., Kleinbeck, S., Juran,S., Blaszkewicz, M., Fricke, H.H., Altmann, L., Berresheim, H.,Bruning, T., 2006a. From chemosensory thresholds to wholebody exposures-experimental approaches evaluating chemosen-sory effects of chemicals. Int. Arch. Occup. Environ. Health 79,308–321.
van Thriel, C., Triebig, G., Bolt, H.M., 2006b. Editorial: evaluationof chemosensory effects due to occupational exposures. Int. Arch.Occup. Environ. Health 79, 265–267.
Walker, J.C., Kendal-Reed, M., Utell, M.J., Cain, W.S., 2001. Humanbreathing and eye blink rate responses to airborne chemicals. Env-iron. Health Perspect. 109 (Suppl. 4), 507–512.
Wise, P.M., Radil, T., Wysocki, C.J., 2004. Temporal integration innasal lateralization and nasal detection of carbon dioxide. Chem.Senses 29, 137–142.
Wise, P.M., Canty, T.M., Wysocki, C.J., 2005. Temporal integration
of nasal irritation from ammonia at threshold and supra-thresholdlevels. Toxicol. Sci. 87, 223–231.Wysocki, C.J., Cowart, B.J., Radil, T., 2003. Nasal trigeminalchemosensitivity across the adult life span. Percept. Psychophys.65, 115–122.