We used aerosol boluses to study convective gas mixing in the lung of four healthy subjects on the ground (1 G) and during short periods of microgravity (μG) and hypergravity (∼1.6 G). Boluses of 0.5-, 1-, and 2-μm-diameter particles were inhaled at different points in an inspiration from residual volume to 1 liter above functional residual capacity. The volume of air inhaled after the bolus [the penetration volume (Vp)] ranged from 150 to 1,500 ml. Aerosol concentration and flow rate were continuously measured at the mouth. The dispersion, deposition, and position of the bolus in the expired gas were calculated from these data. For each particle size, both bolus dispersion and deposition increased with Vp and were gravity dependent, with the largest dispersion and deposition occurring for the largest G level. Whereas intrinsic particle motions (diffusion, sedimentation, inertia) did not influence dispersion at shallow depths, we found that sedimentation significantly affected dispersion in the distal part of the lung (Vp >500 ml). For 0.5-μm-diameter particles for which sedimentation velocity is low, the differences between dispersion in μG and 1 G likely reflect the differences in gravitational convective inhomogeneity of ventilation between μG and 1 G.
- convective mixing
- aerosol bolus
- ventilation inhomogeneity
during the last decade, the aerosol bolus technique has been widely used to characterize convective gas transport in the human lung (1, 3, 4, 9, 14). This technique consists of inserting a small amount of aerosol at a predetermined point in the subject’s inspiratory volume and analyzing the distribution of the aerosol in the subsequent expiration. During its transport through the lung, the aerosol bolus is subject to both dispersion and deposition. Aerosol dispersion is determined by intrinsic motions of the particles resulting from diffusion, sedimentation, and inertia and also by convection. Aerosol deposition is determined by inertial impaction, sedimentation, and diffusion. Particles with a diameter ∼1 μm are usually used in the bolus technique because the intrinsic motions resulting from their inertial, gravitational, and diffusive transport are low. Therefore, these particles are well suited to follow air streamlines and trace convective airflow patterns. The increase in the width of the bolus induced by the passage through the lungs over a respiratory cycle, i.e., the aerosol bolus dispersion, is a direct measurement of the mixing undergone by the particles. This mixing is usually called convective mixing and refers to all the mechanisms that transfer particles from the inhaled air into the resident gas, except Brownian diffusion. Factors contributing to convective mixing include velocity profiles within the air spaces, airway and alveolar geometries, asymmetries between inspiratory and expiratory flows, nonhomogeneous ventilation of the lung, and cardiogenic mixing.
In a previous study (7), we performed bolus inhalations in normal subjects using 1-μm-diameter particles in normal gravity (1 G), in short periods of microgravity (μG), and in hypergravity (∼1.6 G). The results showed that the aerosol bolus dispersion was strongly gravity dependent, with the largest dispersion occurring for the largest G level. In the present study, we report bolus inhalations of 0.5-, 1-, and 2-μm aerosols in 1 G, μG, and in hypergravity. We discuss the effect of the G level on convective mixing as well as how particle size affects aerosol bolus dispersion. The results show that, whereas the intrinsic motions of the particles did not affect dispersion at shallow depths, sedimentation significantly influenced dispersion in the distal part of the lung.
Aerosol bolus data were collected by using the same equipment as used in the previous study (7). A schematic representation of the setup is shown in Fig. 1. Briefly, the setup allowed the injection of an aerosol bolus of ∼70 ml at a given point in the inhaled air by switching computer-controlled pneumatic valves. The measurement of the aerosol concentration and the flow rate were provided by a photometer (model 993000, PARI) (19) and a pneumotachograph (Fleisch no. 1, OEM Medical), respectively. The measurement of the flow rate was not affected by the presence of particles in the breathing air. The photometer, the pneumotachograph, and the valves were heated to body temperature to prevent water condensation. A diffusion dryer was located between the photometer and the mouthpiece. It removed the water vapor from the exhaled air to avoid condensation on the lenses of the photometer.
The bolus tube was filled with aerosol containing monodisperse polystyrene latex particles (Duke Scientific). The particles were supplied in suspension (water), and the concentrate was diluted and dispensed via two Acorn II nebulizers (Marquest Medical Products). Before entering the bolus tube, the aerosol flows through a heated hose and a diffusion dryer to remove water droplets. The sizes of the spherical particles as specified by the manufacturer were 0.497 ± 0.0094, 1.07 ± 0.014, and 2.04 ± 0.044 (SD) μm. For convenience, these are referred to as 0.5-, 1-, and 2-μm-diameter particles, respectively.
The aerosol generated by the nebulizers was checked with a particle sizer (PCS-2000 Special, PALAS). The size analysis confirmed the size of the particles given by the manufacturer and shows than the amount of doublets in the aerosol was <3% for 1- and 2-μm particles and <4.5% for 0.5-μm particles. Aerosol concentration was ∼104particles/cm3 for 0.5- and 1-μm particles and ∼5 × 103particles/cm3 for 2-μm particles. Tests using conductive tubing showed no significant effect from possible electrostatic charges on the particles.
Data Recording and Analysis
A personal computer (IBM ThinkPad 360 CSE) equipped with a 12-bit analog-to-digital card (National Instrument, DAQ700) was used for data acquisition. Signals from the photometer, a G sensor, and the pneumotachograph were sampled at 100 Hz. We used the same custom software for the data acquisition as in the previous study (7).
Data were collected on the ground and aboard the National Aeronautics and Space Administration (NASA) Microgravity Research Aircraft. A typical flight consisted of a climb to an altitude of ∼10,000 m, with the cabin pressurized to ∼600 Torr. A “roller coaster” flight profile was then performed. The aircraft was pitched up at ∼1.6 head-to-foot acceleration (Gz) to a 45° nose-high attitude. Then the nose was lowered to abolish wing lift, and thrust was reduced to balance drag (thus maintaining μG). A ballistic flight profile resulted and was maintained until the aircraft nose was 45° below the horizon. In this manner, μG was maintained for ∼27 s. A pullout averaging ∼1.6 Gz was maintained for ∼40 s, causing the nose to pitch up to a 45° nose-high attitude, and allowed the cycle to be repeated. At the beginning of the pullout phase, the G level varies between ∼1.2 Gz and 1.8 Gz before stabilizing at ∼1.6 Gz. Tests in hypergravity were performed during the stable part of the pullout phase. For convenience, the ∼1.6 G hypergravity phase will be referred to as 1.6 G in the text.
Subjects and Protocol
Four healthy subjects participated in the study. These are the same four subjects who participated in the previous studies (5, 7), and we retain their subject number for comparison purposes. Their relevant anthropometric data are listed in Table 1. After a few normal breaths, the subject exhaled to residual volume (RV) to ensure a known lung volume starting point. As functional residual capacity (FRC) varies significantly with G level (8, 11), we chose to use the more stable RV as the starting point for the test breath. Although there is some small change in RV in μG (8), the effect on our measurements is small. The test breath consisted of an inspiration from RV to FRC + 1 liter at a flow rate of ∼0.45 l/s, immediately followed by an expiration to RV, also at a flow rate of ∼0.45 l/s. A flowmeter provided visual feedback to the subject. FRC refers to the seated 1-G FRC of the subject and was fixed for all experiments on that subject. During the inspiration, an aerosol bolus of ∼70 ml was introduced at different penetration volumes (Vp). The Vp was defined as the volume of air inhaled from the mode of the aerosol bolus to the end of the inhalation. Vps of 150, 300, 500, 800, 1,000, 1,200, and 1,500 ml were used for all three particle sizes, with the exception of 300 ml for 0.5 μm, 1,000 ml for 0.5 and 2 μm, and 1,500 ml for 2 μm.
The protocol was repeated four times for each Vp and each particle size both in μG and 1.6 G. Before the flight, a set of data was also collected on the ground (1 G) with the same protocol. The protocol was approved by both the Committee on Investigations Involving Human Subjects at the University of California, San Diego, and by the Institutional Review Board at the NASA Johnson Space Center, Houston, TX.
For each bolus test, we calculated the aerosol dispersion (H), the aerosol deposition (DE) and the mode shift (MS).
On a graph of aerosol concentration as a function of the respired volume, the half-width is defined as the bolus width (in ml) between the two points of one-half the maximum concentration of the bolus. The change in half-width H reflects the aerosol dispersion and was obtained by the following equation Equation 1whereH in andH ex are the half-width of the inspired and expired boluses, respectively.
Deposition was calculated by using the following equation Equation 2 whereN in andN ex are the numbers of particles in the inspired and expired bolus, respectively.N in andN ex were calculated from the integration of the aerosol concentration as a function of the respired air. The integration was only done when the concentration exceeded 5% of the maximal expired concentration to reduce error due to the noise of the signal. The 5% cutoff was chosen based on the signal-to-noise ratio of the aerosol tracings obtained aboard the aircraft, which was lower than the signal-to-noise ratio of the 1-G tracings.
The MS was defined as the difference between the position of the peak of the expired bolus (Mex) and the volume penetration of the inspired bolus Equation 3 A negative value of MS indicates that the position of the mode of the expired bolus has shifted to a smaller lung volume than the position of the inspired bolus, i.e., that the bolus has moved toward the mouth. The 10–90% time response of the photometer was 70 ms. This response time was short enough that no correction on the position of the mode of the bolus was required.
Statistical analysis was performed by using Systat V 5.0 (Systat, Evanston, IL). Measurements performed for the same experimental conditions with the same subject were not averaged before the statistical analysis was performed. Data were grouped in different categorial variables such as G level (μG, 1 G, and 1.6 G), particle size (0.5, 1, and 2 μm), Vp(150, 300, 500, 800, 1,200, and 1,500 ml), and subject number. A two-way analysis of variance was performed to test for differences between the chosen categorial variables. Post hoc testing by using Bonferroni adjustment was performed for tests showing significantF-ratios. Significant differences were accepted at the P < 0.05 level.
As deposition increases with increasing Vp, increasing particle size, and increasing G level, some tests had such high depositions that reliable measurements of dispersion and mode shift were not possible. For 1-μm-diameter particles, data were discarded for Vp >800 ml at 1.6 G. For 2-μm-diameter particles, data were discarded for Vp >800 ml at 1 G and for Vp >300 ml at 1.6 G. The data for 1-μm-diameter particles are the same as those presented in Ref. 7.
Effect of Particle Size
Dispersion. Figure2,A-C, displays the aerosol dispersion H as a function of Vp for each particle size in μG, 1 G, and 1.6 G, respectively. The data are averaged over the four subjects (means ± SD). At each G level and for each particle size, dispersion increased with increasing Vp. In μG, there were no significant differences in dispersion between particle size, except for Vp = 1,200 ml, where the dispersion of 2-μm bolus (H 2 μm) was significantly smaller (P < 0.05) than dispersion of both 0.5- and 1-μm-diameter aerosol boluses (H 0.5 μm and H 1 μm, respectively) (Fig. 2 A). In 1 G, for Vp >500 ml, there were significant differences in dispersion between particle size. At Vp = 500 ml,H 2 μm was significantly higher than bothH 0.5 μm and H 1 μm(Fig. 2 B). At 1.6 G, no significant differences were found in dispersion between particle size for each Vp, except between 1- and 2-μm-diameter particles at Vp = 300 ml (Fig. 2 C), although the data collected at 1.6 G showed much more scatter that at lower G levels.
Deposition. The effect of particle size on the deposition of the aerosol bolus is shown in Fig.3,A-C, in μG, 1 G, and 1.6 G, respectively, in the same format as for Fig. 2. At each G level and for each particle size, deposition increased with increasing Vp. In μG, the data were not significantly different from one particle size to the other (Fig.3 A). In 1 G and 1.6 G, deposition of 2-μm-diameter particles was significantly higher (P < 0.05) than deposition of both 0.5- and 1-μm-diameter particles for all Vps (Fig. 3,B andC). Significant differences between deposition of 0.5- and 1-μm-diameter particles were also found for Vp >300 ml, both in 1 G and 1.6 G.
Mode shift. Figure4, A-C, shows the effect of particle size on the mode shift in μG, 1 G, and 1.6 G, respectively. At each G level and for each particle size, mode shift became more negative with increasing Vp. In μG, there were no significant differences in mode shift between particle size for each Vp. In 1 G and 1.6 G, there were significant differences in mode shift between particle size for Vp >300 ml. Mode shift became more negative with increasing particle size, i.e., the position of the mode of the expired bolus moved toward the mouth.
Effect of G Level
The effect of G level on aerosol dispersion, deposition, and mode shift for 0.5- and 2-μm-diameter particles is qualitatively similar to the effect of G level on 1-μm-diameter particles that has been extensively described in a previous paper by our group (7), except for mode shift of 0.5-μm-diameter particles at Vp = 1,200 ml. For each particle size, both deposition and dispersion increased with Vp and were gravity dependent, with the greatest dispersion and deposition occurring for the largest G level. At each G level and for each particle size, mode shift became increasingly more negative with increasing Vp (Fig. 4). Table2 summarizes the results of the statistical analysis testing for differences between the data. For each particle size, significant differences between μG and 1-G data and between 1-G and 1.6-G data are shown for dispersion, deposition, and mode shift.
In this study, we report the effect of both G level and particle size on the dispersion and deposition of aerosol boluses inhaled to various volumetric depths within the lung. The measurement of the aerosol bolus dispersion is a means of probing convective mixing at different depths within the lung. Particles have negligible diffusive properties compared with gases and are, therefore, well suited to trace convective gas transport. However, whereas particle intrinsic motions (sedimentation, diffusion, and inertia) are negligible compared with gas diffusive properties, they significantly vary from one particle size to the other. Table 3 summarizes the intrinsic motions of the particles as a function of their size. Particle sedimentation is proportional to the G level and is, therefore, absent in μG. The inertia of the particles is measured by their stopping distance (10). When the direction of the gas flow changes, the suspended particles continue to move in the original direction of flow until they lose momentum as a result of friction with the molecules in the surrounding medium. The stopping distance is defined as the distance traveled by the particles until they follow the new direction of the flow. The stopping distance is a function of the air velocity within the airways. It has been calculated with the same method as used by Hickey (10), using airway dimensions based on Weibel model A (18) and assuming a mouth flow rate of 0.45 l/s. The stopping distance has been calculated for particles located in the trachea and at a volume lung depth of 1,200 ml. Between 0.5- and 2-μm particles, displacements by diffusion varies by a factor of ∼2.2, settling velocity by a factor of ∼13, and particle inertia as inferred from the stopping distance by a factor of ∼15.
The data presented in this study allow us to investigate to which extent differences in intrinsic particle motions affect aerosol bolus dispersion. Because the bolus experiments were performed with the same breathing conditions for each particle size, differences in dispersion at a given Vp should be explicable by differences in intrinsic particle motions and G level. In μG, dispersion was similar for the three particle sizes, except at Vp = 1,200 ml, whereH 2 μm was smaller thanH 0.5 μm and H 1 μm(Fig. 2 A). Sedimentation is absent in μG, and at Vp = 1,200 ml, particle inertia is negligible (Table 3). Therefore, the only intrinsic motion that can be responsible for dispersion is diffusion. Diffusion of 2-μm-diameter particles is low compared with 0.5- and 1-μm particles. Diffusion is more effective in the distal part of the lung where gas velocities are low and, therefore, residence time is high. This may explain the slightly lower dispersion observed deep in the lung for 2-μm particles compared with 0.5- and 1-μm particles.H 2 μm is, however, ∼12% lower thanH 0.5 μmand H 1 μmat Vp = 1,200 ml, suggesting that particle diffusion does play a role in dispersion, although its effect is small.
In 1 G, dispersion increased with both particle size and Vp. At shallow Vps, however, dispersion was similar for the three particle sizes (Fig. 2), suggesting that the effect of the intrinsic particle motions is negligible and that dispersion is mainly attributed to convective mixing at shallow lung depths. This is in agreement with the results of Scheuch and Stahlhofen (15), who reported no significant differences between the dispersion of 0.9- and 1.9-μm-diameter aerosol boluses at Vp = 250 ml in 1 G.
The difference between dispersion in μG and 1 G for each particle size reflects both the increase in ventilatory inhomogeneities from μG to 1 G and the effect of sedimentation on aerosol dispersion. The increase in mixing can be characterized by the change in the slope of the regression lines of dispersion as a function of Vp. The slopes are 0.19 and 0.36 (ml/ml) in μG and 1 G for 0.5-μm particles, 0.20 and 0.52 for 1-μm particles, and 0.16 and 0.70 for 2-μm particles. The slopes for each particle size are the same in μG but differ significantly from one particle size to the other in 1 G. The increase in the slope between particle sizes at 1 G suggests that the sedimentation of the particles is an important factor in the dispersion we measured. On the other hand, in 1 G, although the distance covered in 1 s by 0.5-μm particles due to gravitational sedimentation is low compared with the distance covered by diffusion in 1 s (Table 3), the slope almost doubled between μG and 1 G. This increase in the slope cannot be explained only by the action of gravitational sedimentation but indicates most probably the effect of the gravitational convective ventilatory inhomogeneity on aerosol bolus dispersion.
In a theoretical study, Rosenthal et al. (13) showed that ventilation inhomogeneities induce dispersion. Such ventilation inhomogeneities are dependent on the gravity level. Verbanck et al. (17) performed multiple-breath washout tests in μG and on Earth and showed that, whereas ventilatory inhomogeneities are reduced in μG compared with 1 G, the gravity-independent ventilatory inhomogeneity is at least as large as the gravity-dependent inhomogeneity. In a study of single-breath wash-in tests of helium and sulfur hexafluoride in sustained μG, Prisk et al. (12) also showed the presence of both gravitational and nongravitational ventilatory inhomogeneities. The gravity-independent ventilatory inhomogeneity includes both convective and diffusive effects that are difficult to distinguish when gases are used. When particles are used, diffusive processes are very small. The increase in dispersion as a function of Vp in μG (Fig.2 A) illustrates the presence of convective ventilatory inhomogeneity that is not gravitational in origin. If we consider the 0.5-μm-diameter particles for which both diffusion and sedimentation are small, the difference in dispersion between μG and 1 G reflects mainly the effect of convective ventilatory inhomogeneity that is gravity dependent.
In 1.6 G, differences in dispersion between particle size are reduced compared with dispersion in 1 G. This is perhaps unexpected, because the 1-G data suggest that an increase in particle size, which increases sedimentation rate, will increase dispersion. A possible mechanism is that, as a result of sedimentation, particles carried by one flow streamline during inspiration might be carried by another streamline during expiration, contributing to the increase in dispersion that is measured. On the other hand, in 1.6 G, the higher sedimentation rate than in 1 G causes more particles to deposit (Fig.3 A). The particles that penetrate deeper in the lung deposit more, eroding the distal tail of the bolus and potentially reducing the half-width of the exhaled bolus. The removal of the particles from the bolus tail might explain the smaller dispersion we measured. Whereas our limited data at 1.6 G prevent us from drawing firm conclusions, they are, however, supported by a theoretical analysis of Rosenthal et al. (14), who concluded that deposition is likely to decrease dispersion. Brown et al. (4), in a study on aerosol dispersion in the human lung, also suggested that deposition may be suppressing dispersion. Scheuch and Stahlhofen (15) studied the effect of deposition and dispersion of aerosols in human subjects. They performed tests with various breath-holding periods after the inhalation of an aerosol bolus and found a similar effect of sedimentation on aerosol dispersion to what our data suggest. They found that, for aerosol particles >2 μm, bolus dispersion increased with increasing periods of breath holding. After reaching a maximum, they found that dispersion decreased with even longer breath hold, probably because of the increase of particle loss to the airway walls. Similarly, our data in 1 G show an increase in dispersion with increasing particle size (increasing sedimentation rate). When the sedimentation rate was increased even more by performing the experiments in 1.6 G, dispersion was reduced.
We also compared our 1-G data with a study of Schulz et al. (16) on the influence of intrinsic particle properties on aerosol bolus dispersion. They performed bolus inhalation experiments in dogs with 0.5-, 1-, and 2-μm-diameter particles. They found thatH 1 μm andH 0.5 μmwere similar in the dead space and thatH 1 μm was slightly higher thanH 0.5 μmbeyond it. Similarly,H 2 μm was not significantly different fromH 0.5 μmin the dead space but significantly different beyond it. Although these observations were made in dog lungs, they qualitatively agree with our observations in 1 G.
In μG, no significant differences were found in deposition between particle sizes (Fig. 3 A). This is in sharp contrast to the situation in 1 G and 1.6 G, where deposition increased both with particle size and with G level, illustrating the effect of sedimentation on deposition. This observation is consistent with our previous study of total aerosol deposition in μG, which showed no size dependence in deposition over this range of particle sizes (6). If inertial impaction and Brownian diffusion were the main causes of deposition in μG (sedimentation being absent), we would expect different deposition profiles as a function of Vp for the three particle sizes. For the smallest particle size, we would expect low deposition by inertial impaction and, therefore, lower deposition in the upper airways than for larger particle sizes. Conversely, there should be higher deposition deeper in the lung because of Brownian diffusion for the smallest particles.
In 1 G and 1.6 G, differences in deposition between particle size were small at shallow Vps and increased with increasing Vp, with the highest deposition occurring for the largest particle size. Comparison between deposition in μG and 1 G strongly shows that among the different intrinsic motions sedimentation is the one that affects deposition the most in 1 G for this range of particle sizes.
We also compared our 1-G results with those obtained by Schulz et al. (16) in dog lungs. They found that deposition at shallow Vps was similar for 0.5- and 1-μm particles and that at larger Vps deposition of 1-μm particles was higher than deposition of 0.5-μm particles. For 2-μm particles, deposition was already larger at shallow Vps, compared with the deposition of 0.5-μm particles. These data qualitatively agree with ours.
At shallow Vps, the mode shift for each G level and particle size was almost zero. At larger Vps, the mode of the expired boluses was shifted to smaller volumes, i.e., toward the mouth. The shift might be explained both by an asymmetry of lung filling and emptying and by particle deposition that alters the shape of the expired boluses.
Mode shift behaved similarly to dispersion and deposition in both μG and 1 G. In μG, mode shift was almost similar for each particle size at any given Vp. Although there was no significant difference between particle size, at large Vp, there was a trend for mode shift to become smaller (closer to zero) with increasing particle size. The trend was reversed in 1 G and 1.6 G, where the mode shift became larger (more negative) with increasing particle size. At a given G level, ventilatory inhomogeneity does not vary, and differences in mode shift should be attributed only to differences in particle deposition. Indeed, at each G level, mode shift appeared to qualitatively vary in the same way as deposition (compare Figs. 3 and 4).
In summary, aerosol bolus inhalations were performed with 0.5-, 1-, and 2-μm-diameter particles in four subjects on the ground and during short periods of weightlessness and hypergravity. The boluses were inhaled at different Vp values, ranging from 150 to 1,500 ml. The data showed that, for each particle size, both bolus dispersion and deposition increased with Vp and that they were gravity dependent, with the largest dispersion and deposition occurring for the largest G level. In the particle size range we studied, we showed that, among the intrinsic motions of the particles, sedimentation was the mechanism that had the greatest effect on both dispersion and deposition. The data also permitted us to distinguish between gravitational and nongravitational convective ventilatory inhomogeneities. The data suggested that gravitational convective inhomogeneity affects dispersion in the distal part of the lung. In μG, the increase in dispersion with Vp showed considerable nongravitational convective inhomogeneity in the normal human lung. These observations are in agreement with previous studies using gases that also showed the presence of similar inhomogeneity.
The authors acknowledge the collaboration of Janelle Fine, Jeff Struthers, Bob Williams, and Noel Skinner and the administrative assistance of Mary Murrell and Marsha Dodds. They also thank Manuel Paiva, Christa Roth, and Joachim Heyder for scientific and technical support and Sylvia Verbanck from the Vrije Universiteit van Brussel (Belgium) where the size analysis of the aerosols was performed.
Address for reprint requests and correspondence: C. Darquenne, Physiology/NASA Laboratory 0931, Dept. of Medicine, University of California, San Diego, 9500 Gilman Dr., La Jolla, CA 92093-0931 (E-mail:).
This work was supported by National Aeronautics and Space Administration Grant NAGW-4372.
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- Copyright © 1999 the American Physiological Society