Acute in vivo pulmonary toxicity assessment of occupationally relevant particulate matter from a cellulose nanofiber board

Abstract

Cellulose nanofibers (CNFs) are an emerging engineered nanomaterial that are utilized in a variety of applications, including as a replacement for urea-formaldehyde, and other adhesives, as the binding agent in manufactured fiber and particle boards. To ensure the health and well-being of those producing, installing, or otherwise using cellulose nanofiber boards (CNFBs) it is imperative that the particulate matter (PM) produced during CNFB manipulation be evaluated for toxicity. We developed and internally verified a generation system to examine the PM produced by sanding CNFB using aluminum oxide sandpaper. With 80-grit sandpaper our system produced a low dispersity aerosol, as determined by a scanning mobility particle sizer and an optical particle counter, with a geometric mean of 28 nm (GSD = 1.60). ICP-MS evaluation showed little difference in metal concentrations between CNFB PM and nonsanded CNFB stock. We then used the system to simultaneously generate and expose both male and female C57BL/6J mice acutely for 4 hours at a concentration of 7.9 mg/m³. Sham-exposed controls were treated similarly but without sanding the CNFB. Analysis of bronchoalveolar lavage (BAL) fluid biomarkers showed no signs of inflammatory response at either 4- or 24-hours post exposure. Further, BAL cell viability, number of total cells, and pulmonary cellular recruitment were not significantly changed between the sham-exposed controls and CNFB-exposed mice. Histology further confirmed no pulmonary toxicity as a result of CNFB PM inhalation. We conclude that inhalation of a high concentration of the PM from manipulation of a CNFB did not produce acute toxic responses within 24 hours of exposure.

Fig. 1.

Diagram of the sanding aerosol generation and exposure system (SAGES) in conjunction with a pair of nose-only inhalation towers and auxiliary characterization devices. Design components and operation of the SAGES are discussed in detail in section 2.1. Red tubing indicates particle-laden airflow. Magnification of an exposure tower, outlined by the green dashed box, displays a more detailed representation of the nose-only exposure design. Calibrated pDRs were used to measure real time aerosol concentrations and determine a TWA exposure concentration. The combined use of a SMPS and an OPC allowed for aerosol size characterization for particles ranging from 10 nm to 30 µm.

Fig. 2.

SAGES generated particle size distributions of aerosols produced from medium density fiber board using aluminum oxide sanding belts of various coarseness. Samples of fiber board were sanded at a feed rate of 0.5 mm/min. Size distributions were determined through the combined use of a scanning mobility particle sizer and an aerosol spectrometer/optical particle counter. The resulting data from the instruments were corrected for bin width and dN/dlogD_p values were assigned to the bin midpoint before separate instrument data were combined for complete size distribution comparison. The inset shows the ultrafine size distribution of p320 and p400 grits on a different Y-axis scale.

Fig. 3.

(a) Concentration vs. time plot of CNFB aerosol within the exposure towers of two CNFB stocks of different surface areas as monitored by a calibrated pDR. The spikes in concentration occurred during sampling for size distribution analysis by SMPS and OPC. Dashed lines through each concentration profile indicate the TWA concentrations of 13.5 and 5.4 mg/m³ that were determined gravimetrically from the pDR filters. (b) Aerosol size distributions produced from two CNFB stocks of different surface areas as determined by SMPS and OPC. Shaded areas around mean lines display the standard deviation of the eight samples taken during each experiment. The aerosols had GMs (GSDs) of 27 (1.52) and 24 nm (1.49) for the 507 and 273 mm² CNFB stocks, respectively.

Fig. 4.

(a) Tower concentration vs. exposure duration of the CNFB and sham exposures as monitored by gravimetrically calibrated pDRs. The dashed line indicates the TWA total particulate concentration over the duration of the CNFB exposure (7.9 mg/m³) determined gravimetrically from pDR filters. The dotted line indicates the TWA CNFB mouse inhalable fraction (4.1 mg/m³. The SAGES control exposure had a TWA concentration of 0.22 mg/m³ determined from pDR filter gravimetric analysis. (b) Aerosol size distributions of the CNFB and sham exposures. The CNFB and sham exposures had a GM(GSD)s of 28 nm (1.48) and 39 nm (2.24), respectively. Gray area surrounding average line shows standard deviation of the size distribution samples from each exposure. (c and d) Aerosol temperature and relative humidity over the exposure duration.

Fig. 5.

(a) Total BAL cells per mouse compared by treatment and latency grouping. (b) BAL cell viability compared by treatment and latency grouping. Bars indicate mean and standard deviation. One-way ANOVA testing revealed lack of statistically significance differences between sex- and latency-matched sham and exposure groups.

Fig. 6.

(a) LDH concentration within BAL supernatant compared by treatment and latency grouping. (b) Total protein concentration with BAL supernatant compared by treatment and latency grouping. Bars indicate mean and standard deviation. One-way ANOVA testing revealed lack of statistically significance differences between sex- and latency-matched sham and exposure groups.