This period was followed by a 60 min exposure period and a 30 min

This period was followed by a 60 min exposure period and a 30 min recovery period. From RAD001 mw the baseline period, the mean value for each parameter was calculated for each animal. These values were used as the ‘baseline’ (‘control’) values (made equal to 100%) to calculate increases or decreases during exposure and recovery periods. The Notocord

Hem (Notocord System SA, France) data acquisition software was used to collect respiratory parameters. Modules and settings for data acquisition and calculations were as previously described as were the head-out body plethysmographs, pneumotachographs, transducers, and transducer signal amplifiers (Larsen et al., 2004). For each terpene reaction product, the combined exposure-effect was evaluated from the effect on the respiratory frequency that may be decreased by either TB and/or TP elongations, and/or airflow limitation or a combination. However, an evaluation of the specific parameters for sensory irritation, VT, airflow limitation, and pulmonary irritation is necessary to characterize each of the specific airway effects. Further, rapid shallow breathing is another type of pulmonary irritation which decreases TP and VT, increases the respiratory frequency and decreases TI. However, neither increase in respiratory frequency nor decrease in TI was observed. Thus, only TP elongations were evaluated. The effects may have different time-dependent relationships; SAHA HDAC thus, a single effect

may dominate in one period and may overlap (coincide) other effects in other exposure periods. An exposure-dependent effect was considered reversible if it within its recovery period normalized or nearly normalized to the pre-exposure value and exposures reached approximately the same level as the lower concentrations. Time–response relationships for the decrease in respiratory frequency and airflow limitation and the increase in TB were plotted as 1-min mean values of the groups. Linear log concentration response relationships were used to establish concentration-effect relationships for the airway effects by means of MS Excel. The depression in respiratory frequency (RD) as percentage

of the pre-exposure baseline level was used as endpoint to determine the no-observed-(adverse)-effect level (NO(A)EL) of the reflex-mediated response in mice (RD0). The regression line was used for estimating the concentration that depressed the respiratory frequency Chlormezanone by 0% in the exposure period 11–20 min where it had its maximum. The NOEL for sensory irritation was also obtained from TB by regression; the threshold of increase in TB elongation was obtained by extrapolation to 100% of the preexposure level (TB100). Furthermore, the NOEL for airflow limitation was estimated from the mean effect at the exposure period from 46 to 60 min. The value was obtained by extrapolation of VD/VT to the preexposure level of 100% ((VD/VT)100). All NOELs are given together with their respective 95% confidence interval in Section 3.

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