Quantification of cigarette smoke particle deposition in vitro using a triplicate quartz crystal microbalance exposure chamber

Abstract

There are a variety of smoke exposure systems available to the tobacco industry and respiratory toxicology research groups, each with their own way of diluting/delivering smoke to cell cultures. Thus a simple technique to measure dose in vitro needs to be utilised. Dosimetry-assessment of dose-is a key element in linking the biological effects of smoke generated by various exposure systems. Microbalance technology is presented as a dosimetry tool and a way of measuring whole smoke dose. Described here is a new tool to quantify diluted smoke particulate deposition in vitro. The triplicate quartz crystal microbalance (QCM) chamber measured real-time deposition of smoke at a range of dilutions 1:5-1:400 (smoke:air). Mass was read in triplicate by 3 identical QCMs installed into one in vitro exposure chamber, each in the location in which a cell culture would be exposed to smoke at the air-liquid interface. This resulted in quantification of deposited particulate matter in the range 0.21-28.00 μ g/cm(2). Results demonstrated that the QCM could discriminate mass between dilutions and was able to give information of regional deposition where cell cultures would usually be exposed within the chamber. Our aim is to use the QCM to support the preclinical (in vitro) evaluation of tobacco products.

Figure 1

British American Tobacco's standard exposure chamber used for in vitro exposures to whole smoke at the air-liquid interface was modified to accommodate the 3 QCM units. The picture shows a top view of the 3-in-1 QCM exposure chamber base (crystal ø = 2.54 cm; cell support insert ø = 2.4 cm). QCM position 1 is proximal to the passive exhaust port ∗. QCM positions 2 and 3 are distal to the exhaust port and behind position 1 on the right and left, respectively.

Figure 2

A schematic diagram of the triplicate QCM exposure chamber. (a) Crystals are installed directly into and replace the 3 positions where (b) cells would usually be exposed to whole smoke at the air-liquid interface. Illustration by J. Adamson.

Figure 3

Triplicate QCM deposition data. (a) An individual value plot showing deposited particle mass quantification of ISO whole smoke (9.4 mg) diluted in the range 1 : 5–1 : 400 (smoke : air, v/v), n = 5/position. (b) A multi-vari chart of mean deposited mass at all five dilutions 1 : 5–1 : 400. The chart shows the average distribution of particulate deposition around the chamber within the range tested, from 25 values (5 dilutions at n = 5/dilution). There was no statistically significant difference in deposition between the three QCM positions.

Figure 4

An interaction plot of the data means for the single unit QCM [15] and the 3-in-1 QCM, at the five airflows tested. There was no statistically significant difference between the two different devices.

Figure 5

Deposition when a Cambridge filter pad (CFP) was placed prior to the QCM chamber to trap particulate matter. Cigarettes (3R4F) were smoked at the ISO regime at a dilution of 1 : 5 (smoke : air, v/v) for 1 hour. When the scale is adjusted to see the trace more clearly, note the distinct and repeated “n-shaped” pattern of mass increase and decrease per puff over time. The 3 traces represent the 3 QCMs within the chamber: QCM position 1, bottom; QCM position 2, top; QCM position 3, middle.