Here, we establish that aerosolization of a commercial e-liquid by a pneumatic atomizer at 200 °C provides a metal-reduced aerosol that has aldehyde levels comparable to those found in real-world e-cigarette aerosol.
Research Article
Generation of Substantially Metal-Reduced but Aldehyde-Rich Aerosol for E-Cigarette Toxicity Studies
In This Article
Summary
Abstract
When e-liquid is aerosolized in an e-cigarette for inhalation by the user, the aerosol becomes contaminated by a mixture of toxicants, including metals and aldehydes. A method to generate e-cigarette aerosol with high aldehyde but low metal content is lacking, yet is needed to untangle the potential independent effects versus toxicological interactions of aldehyde and metal exposure from e-cigarette aerosol mixtures.
The objective of this study was to establish a method for generating aldehyde-rich aerosols from e-liquids that lack, however, the large metal contamination typically found in e-cigarette aerosol. To aerosolize e-liquid at temperatures that promote aldehyde formation (up to 200 °C), a pneumatic atomizer was externally heated. Both a pure propylene glycol/vegetable glycerin (PG/VG) mixture and a commercial e-liquid were aerosolized. Aerosols were analyzed for known e-cigarette-associated toxicants, including 11 aldehydes using liquid chromatography-tandem mass spectrometry and 9 metals (Cr, Cu, Fe, Mn, Ni, Pb, Sr, V, Zn) using inductively-coupled plasma mass spectrometry.
Aldehyde levels in aerosols from the pneumatic atomizer only reached those in a real-world e-cigarette aerosol if the commercial e-liquid was aerosolized at 200 °C. Metal levels in the aerosols from the pneumatic atomizer were then at least 8 times lower than in e-cigarette aerosol from a "Mod" device operated at 50 W. Cu, Ni, and Pb levels were particularly low across all aerosol samples, with some levels below the method detection limit (MDL).
Aerosolization of a commercial e-liquid by a pneumatic atomizer at 200 °C produces aerosol that is low in metal levels (particularly Ni, Cu, Pb), but has aldehyde levels comparable to those found in a real-world e-cigarette aerosol.
Introduction
In the US, e-cigarettes are the most commonly used tobacco product among youth; in 2024, 5.9% of students reported using e-cigarettes1. This is a public health concern because e-cigarette aerosols, which are inhaled by e-cigarette users, contain a broad range of toxic chemicals that, upon inhalation, can have various adverse health effects2. Some toxic chemicals in aerosols may not only emanate from the original e-liquid formulation but also be introduced during device use. For instance, metals leach from the metallic heating coil used to aerosolize the e-liquid through the process of dissolution and evaporation into the aerosol3. In contrast, most of the aldehydes present in e-cigarette aerosol are formed from the base ingredients of e-liquids, propylene glycol (PG) and vegetable glycerin (VG), when the temperature of the e-liquid is increased to the boiling point of the e-liquid for the purpose of aerosolization4.
The presence of metals and aldehydes in e-cigarette aerosol is of particular concern because several of those are toxic, especially to the brain and other organs. For instance, the metals lead (Pb), manganese (Mn), copper (Cu), iron (Fe), nickel (Ni), and chromium (Cr) are known neurotoxicants which have been found to accumulate in the brains of mice exposed to e-cigarette aerosol, at levels previously described to cause neurotoxic effects5. High Mn and Pb exposure, Fe overloading in the brain, and Cu imbalance have all been linked to a higher risk for the development and/or progression of neurodegenerative diseases, e.g., Parkinson's disease (PD). This metal-induced neurotoxicity usually involves cellular mechanisms such as oxidative stress due to reactive oxygen species (ROS) overproduction, mitochondrial dysfunction, protein aggregation, and neuroinflammation6,7,8.
The aldehydes acrolein and formaldehyde have also been linked to neurodegeneration9,10,11, possibly leading to late-onset neurological diseases such as PD. As shown in a PD mouse model, aldehyde build-up can be toxic by promoting protein aggregation12, and individuals with a mutation in the enzyme responsible for the conversion of acetaldehyde into acetic acid (ALDH2) may have a greater risk of Alzheimer's disease13. Even though it is known that metals and aldehydes can be neurotoxic individually, less is known about their effects as mixtures, and it appears quite challenging to untangle the independent and interaction effects of these two chemical groups in the context of e-cigarette aerosol exposure. To properly study chemical class-related toxicological effects, the capability is needed to generate simulated aerosols that are metal-reduced compared to aerosols from a chosen reference e-cigarette while matching its aldehyde levels.
The objective of this paper is to establish a method that, like in an e-cigarette, aerosolizes e-liquid at high temperature to generate an aldehyde-rich aerosol, which, however, lacks substantial metal contamination, to provide the toxicology field with a new tool to untangle the respective toxic effects of these chemicals within e-cigarette aerosol mixtures. To achieve this goal, we avoided aerosolizing the e-liquid by using a metallic heating coil, which would act as a source of metal contamination. Instead, we used a pneumatic atomizer14, which was heated externally to promote the formation of aldehydes from e-liquids. To evaluate our approach, we compared aldehyde and metal levels in aerosols from the atomizer to those from a "Mod" e-cigarette we used to expose rodents to e-cigarette aerosol.
Protocol
Setting up the pneumatic atomizer
All materials used in the study are listed in the Table of Materials file. To generate simulated e-cigarette aerosol without using a metallic heating coil that releases metals, we used a commercial pneumatic atomizer. To achieve the high temperatures needed to promote aldehyde formation, we heated the pneumatic atomizer as a whole by wrapping heating tape around it. To that end, we loosened the two screws, which secure the actual aerosol unit to its stand, such that the heating tape could be guided through the resultant gap instead of wrapping it around the stand. The temperature of the pneumatic atomizer was measured by attaching a platinum resistance temperature detector to its exterior surface, i.e., the temperature of the e-liquid was not directly measured. The thermal connection was improved by applying a small amount of thermal paste to the detector. The measured temperature was read in by a microcontroller board, which controlled a relay to supply the heating tape with 110 V line power if the actual temperature was smaller than a target temperature (between 50 °C and 200 °C). Figure 1 depicts the experimental setup.
Tubing materials that are needed to operate the pneumatic atomizer and that were originally supplied by manufacturer had to be replaced by more temperature resistant materials: 1) the liquid feed tubing (14-gauge PTFE) was replaced by high-temperature silicone tubing (1/8" OD), 2) the high-molecular weight polyethylene tubing sections (1/4" OD) of the excess liquid drain line and the filtered air supply were replaced by extreme-temperature hard plastic tubing (1/4" OD).
Figure 1: Diagram of experimental setup. Please click here to view a larger version of this figure.
Operating the pneumatic atomizer
Operating the pneumatic atomizer in the default recirculation mode could cause metal accumulation in the recirculating e-liquid, which is the excess, non-aerosolized e-liquid that flows through the excess liquid drain-off back into the e-liquid bottle. To prevent associated metal accumulation in the generated aerosol, the pneumatic atomizer was operated in non-recirculation mode, in which excess e-liquid flows into an added waste bottle. House compressed air was filtered and dried before supplying it to the atomizer via a filtered air supply, which consists of a coalescing pre-filter, a coalescing filter, a carbon filter, and a membrane dryer to remove solid particulate matter, oil and water droplets, and oil and water vapor. According to the manufacturer, the final carbon filtration removes 99.99995% of particles > 100 nm in size.
Aerosol collection
The pneumatic atomizer was used to generate aerosols from either a 70%vol PG/30%vol VG solution or a commercial, real-world e-liquid with 6% nicotine content that we used in previous research15. Aldehyde and metal levels of the aerosols produced by the pneumatic atomizer were compared to those in aerosols generated from the commercial e-liquid using a Mod e-cigarette, i.e., a modifiable device which can be refilled with e-liquid and the operating power of which can be adjusted. Recently, this Mod was used for chronic exposure of rodents to e-cigarette aerosol produced at 30 W5. In this study, the device was operated at a power output of either 30 W or 50 W.
In all the experiments, including those with the pneumatic atomizer and those with the Mod e-cigarette, aerosol samples were collected in triplicate using the method developed by Olmedo et al.16 for subsequent metal and aldehyde analyses. Aerosol samples were analyzed for 9 metals (Cr, Cu, Fe, Mn, Ni, Pb, Sr, V, Zn) using inductively-coupled plasma mass spectrometry (ICP-MS) and 11 aldehydes (acetaldehyde, acetone, acrolein, benzaldehyde, 2-butanone, butyraldehyde, crotonaldehyde, formaldehyde, hexaldehyde, propionaldehyde, pentanal) by liquid chromatography tandem mass spectrometry (LC-MS/MS) (see Supplementary File 1 and Supplementary Table 1 and Supplementary Table 2 for details). Potential metal transfer between the collected aerosol and the collecting tubing system was not a concern, except for Zn.
Electrochemical sensor for real-time aldehyde survey
A low-cost electrochemical sensor was used to compare in real time the aerosol aldehyde concentrations produced from different e-liquids at different temperatures, and thus be able to adjust more quickly the conditions that favor the generation of aldehydes as compared to offline analysis by LC-MS/MS. That sensor, according to the manufacturer, measures formaldehyde concentrations with a maximum reading of 5,000 ppb. The sensor could be directly connected to a computer via a USB cable, and formaldehyde concentrations could be viewed in real time using software provided by the sensor manufacturer. These real-time measurements enabled us to identify the optimal conditions for generating aldehydes with our pneumatic atomizer. To protect the sensor from the aerosol, the sensor was placed in a small box that allowed for the exchange of ambient air through a small-pore-size membrane that covered a hole in the box (while blocking aerosol droplets).
Statistical analysis
Box plots were used to visualize the distributions of the measured metal and aldehyde concentrations in aerosol and e-liquid (Figure 2). To get a sense of the overall aldehyde and metal concentration of an aerosol sample, we aggregated the data. We summed up the concentrations of the nine measured metals to determine a "total" metal concentration. Similarly, we summed up the concentrations of the eleven measured aldehydes to determine a "total" aldehyde concentration. All statistical analyses were performed using the R software version 4.3.3. Differences in metal concentrations across different aerosols and e-liquids were analyzed by t-test. When p-values were less than 0.05, differences were considered significantly different.
Results
Exploring the conditions that promote aldehyde formation
Aerosols were generated in four phases. At first, we used the pneumatic atomizer to generate aerosols from the PG/VG solution at 50 °C and 100 °C, and we used the "Mod" e-cigarette to generate aerosol from the commercial e-liquid at 30 W and 50 W operating power. As determined by LC-MS/MS, total aldehyde levels were much lower in the PG/VG aerosols than in real-world e-cigarette aerosol (Figure 2A).
Since it took a few weeks to get these results back from the laboratory, we used the electrochemical sensor to explore conditions that would yield higher aerosol formaldehyde levels. We found that formaldehyde levels measured about 2 cm above the outlet tube of the pneumatic atomizer were about 50 ppb and 160 ppb for 50 °C and 80 °C, respectively. We therefore increased the temperature to 100 °C and 140 °C, because Jaegers et al.17 showed that formaldehydes can be generated from PG and glycerol at 133 °C (even though those experiments were not performed using an e-cigarette). For both temperatures, the formaldehyde sensor maxed out, with a reading of 5,000 ppb. However, at 140 °C the sensor maxed out much more rapidly (within one minute), whereas it took 5 min at 100 °C, with some readings still slightly below 5,000 ppb after maxing out.
Given the measurements with the electrochemical sensor, we decided to generate aerosols from a PG/VG solution at even higher temperatures, because e-liquid aerosolized in e-cigarettes is close to the boiling point, which is on the order of 200 °C and higher, depending on the PG/VG ratio18 and other chemicals added to the PG/VG base. We therefore collected aerosols generated at 140 °C and 180 °C and analyzed them using LC-MS/MS. Aldehyde levels were still much lower (5-fold) in the PG/VG aerosols than in the real-world e-cigarette aerosols that we generated with a Mod device at 30 W (Figure 2A).
In our last attempt to increase aldehyde levels in aerosols from the pneumatic atomizer, we aerosolized the commercial e-liquid (also used to generate aerosol from the Mod device), because flavoring chemicals can promote the formation of aldehydes during e-cigarette use19. We generated aerosols from the commercial e-liquid at 140 °C and 200 °C. After ascertaining that the electrochemical sensor indicated the presence of aldehydes, we collected aerosol samples and analyzed them with LC-MS/MS. For 140 °C, total aerosol aldehyde levels were still much smaller than in real-world e-cigarette aerosols. At 200 °C, however, aldehyde levels were comparable to those generated by the Mod device operated at 30 W and only about 30% lower than for e-cigarette aerosol generated by the Mod device operated at 50 W (see Figure 2A).
Figure 2: Total aldehyde and total metal concentrations in two unused e-liquids (PG/VG and commercial e-liquid). (A) Total aldehyde concentration. (B) Total metal concentration. The aerosols generated from the two e-liquids by the pneumatic atomizer heated to different temperatures (50, 100, 140, 180, 200 °C), and the aerosols generated from the two e-liquids by a Mod e-cigarette used at two operating powers (30 and 50 W). Please click here to view a larger version of this figure.
Summary of aerosol aldehyde levels
In summary, total aldehyde levels in aerosols generated with the pneumatic atomizer were only high and comparable to total aldehyde levels generated by the e-cigarette when the commercial e-liquid was aerosolized at 200 °C. The mean total aldehyde concentration for the commercial e-liquid aerosolized at 200 °C was 3 to 12 times higher than the mean concentration of the other five aerosols generated with the pneumatic atomizer for different temperatures or e-liquids; all differences are statistically significant, except for the comparisons to PG/VG aerosolized at 50 °C (p = 0.21) and 180 °C (p = 0.052). When the commercial e-liquid was aerosolized at a lower temperature of 140 °C, the total aldehyde level in the aerosol was 2.7-fold lower and did not differ (factor of 1.0) from the level in the unused commercial e-liquid. For aerosols that were generated from PG/VG at four different temperatures, aldehyde concentrations were generally lower than the concentrations obtained at 200 °C for the commercial e-liquid, except for one out of the three PG/VG samples aerosolized at 50 °C. Its total aldehyde concentration exceeded all triplicate measurements for the commercial e-liquid aerosolized at 200 °C; however, the mean total aldehyde concentration of the triplicate samples was still much smaller (2.7-fold) than the mean for the commercial e-liquid aerosolized at 200 °C (Supplementary Table 3).
The total aldehyde concentrations in the real-world e-cigarette aerosol were mostly driven by acetaldehyde, propionaldehyde, as well as acrolein and formaldehyde (see Supplementary Figure 1). While propionaldehyde is considered to have relatively low toxicity potential, formaldehyde, acrolein, and acetaldehyde are well known for their neurotoxicity10,20. The presence of acrolein in the real-world e-cigarette aerosols could only be mimicked by the pneumatic atomizer when the commercial e-liquid was aerosolized at 200 °C (Figure 3). All other aerosols from the pneumatic atomizer had substantially and significantly lower acrolein concentrations. The formaldehyde concentrations from the e-cigarette operated at 50 W were clearly higher compared to the e-cigarette operated at 30 W and the aerosols from the pneumatic atomizer; however, this finding was not statistically significant (p = 0.1).
Of note, the levels of the aldehydes acrolein, acetaldehyde, and formaldehyde in the aerosols generated from the commercial e-liquid using both the pneumatic atomizer operated at 200 °C and the e-cigarette are substantially higher than the levels in the unused commercial e-liquid (Figure 3).
Figure 3: Acrolein, acetaldehyde, and formaldehyde concentrations in two unused e-liquids (PG/VG and commercial e-liquid). The aerosols generated from the two e-liquids by the pneumatic atomizer heated to different temperatures (50, 100, 140, 180, 200 °C), and the aerosols generated from the two e-liquids by a Mod e-cigarette used at two operating powers (30 and 50 W). Black and gray border colors of the symbols indicate above and below MDL measurements, respectively. Please click here to view a larger version of this figure.
Summary of aerosol metal levels
The total metal concentrations in aerosols generated with the pneumatic atomizer are lower but on the same order of magnitude as the concentrations of aerosol from the e-cigarette generated at 30 W, but substantially lower than for e-cigarette aerosol generated at 50 W (see Figure 2B). The ratios of the mean total metal concentrations in e-cigarette aerosol generated at 50 W to the mean total metal concentrations in aerosols from the pneumatic atomizer range from 8 to 59, with the two limits corresponding to the commercial e-liquid aerosolized at 200 °C and PG/VG aerosolized at 50 °C, respectively. For e-cigarette aerosol generated at 30 W, these ratios range from 2 to 11. Of note, several of the concentrations of individual metals measured in the aerosols from the pneumatic atomizer are much lower than their corresponding levels in aerosols generated with the e-cigarette (see Supplementary Figure 2) and even below their respective method detection limit (MDL; see Supplementary Table 4).
The total metal concentrations in real-world e-cigarette aerosol are mostly driven by Ni and Zn (see Supplementary Figure 2 and Figure 4). For aerosol generated at 50 W, both the Ni and Zn concentrations exceed 1 mg/g, i.e., one part per thousand. Overall, except for Cr, the concentrations of metals are much lower in aerosols generated by the pneumatic atomizer compared to the aerosol from the e-cigarette. Pb, Cu, and Ni levels are particularly low across all aerosol samples from the pneumatic atomizer, with some levels below the MDL. The mean concentrations of Cu, Mn, Ni, and Pb in aerosols generated from the commercial e-liquid at 200 °C are 128, 8, 144 and 75 times lower than those in aerosol from the e-cigarette operated at 50 W (all differences statistically significant), and 48, 1.5, 18 and 18 times lower compared to 30 W (differences not statistically significant) (see Figure 4). Cr levels, however, are only 1.3 times lower compared to aerosol from the e-cigarette operated at 50 W, and even 2 times higher than for 30 W.
The levels of Cr, Cu, Mn, Ni, and Pb in the aerosols generated from the commercial e-liquid using both the pneumatic atomizer and the e-cigarette were substantially higher (for the 200 °C aerosols 47, 2, 29, 97, and 42 times, respectively) than the concentrations in the unused e-liquid (Figure 4). For Cr, Ni, and Pb, levels in the unused commercial e-liquid were below the MDL, which was not the case for the aerosols generated from the e-liquid. Among the four metals, mean concentrations of Cr and Ni were particularly high (15 ng/g and 21 ng/g) compared to Mn and Pb (7 ng/g and 2 ng/g).
Figure 4: Concentrations of five metals in two unused e-liquids (PG/VG and commercial e-liquid). The aerosols generated from the two e-liquids by the pneumatic atomizer heated to different temperatures (50, 100, 140, 180, 200 °C), and the aerosols generated from the two e-liquids by a Mod e-cigarette used at two operating powers (30 and 50 W). Black and gray border colors of the symbols indicate above and below MDL measurements, respectively. Please click here to view a larger version of this figure.
In conclusion, aerosolization of a commercial e-liquid by a pneumatic atomizer at 200 °C provides a substantially metal-reduced aerosol that has aldehyde levels comparable to those found in real-world e-cigarette aerosol. These aerosols have substantially reduced Pb, Ni, and Cu levels and comparably high acrolein levels when compared to e-cigarette aerosol from the Mod device we used in this study. We have been using these aerosols in recent animal experiments to disentangle the neurotoxic effects of metal and aldehyde exposure due to e-cigarette use.
DATA AVAILABILITY:
All raw data, including a data dictionary, were uploaded as supplementary files (Supplementary File 2).
Supplementary Figure 1: Concentrations of all 11 aldehydes analyzed in e-liquids and aerosols. Please click here to download this file.
Supplementary Figure 2: Concentrations of all nine metals analyzed in e-liquids and aerosols. Please click here to download this file.
Supplementary Table 1: MDLs for metals analyses in ng/g. n: number of samples used for analysis. Please click here to download this file.
Supplementary Table 2: Precursors, quantitative and confirmation ions, de-clustering potential, and collision energy for LC-MS/MS analysis of aldehydes. Please click here to download this file.
Supplementary Table 3: Mean (range) of aldehyde concentrations in units of ng/g measured in each e-liquid or aerosol. All conditions were measured in triplicate except for unused PG/VG and commercial e-liquid that came from a single source and were only measured once. Below MDL measurements were replaced by MDL/√2. Please click here to download this file.
Supplementary Table 4: Mean (range) of metal concentrations in units of ng/g measured in each e-liquid or aerosol. Please click here to download this file.
Supplementary File 1: Details of metal and aldehyde analyses. Please click here to download this file.
Supplementary File 2: Raw data zip folder. Please click here to download this file.
Discussion
The total aldehyde concentration (11 aldehydes) of aerosols we generated with the pneumatic atomizer from commercial e-liquid at a temperature of 200 °C was on the order of those measured in aerosols from a Mod device operated at 30 or 50 W. In particular, acrolein occurred at the high levels observed in the real-world e-cigarette aerosols. However, levels of formaldehyde did not match those observed in the e-cigarette aerosols. Altogether, the levels of aldehydes reported in our study, which range from low ng/g to low µg/g of aerosol (summarized in Supplementary Figure 1), are in line with but for the most part in the lower range of those reported in other studies4,21. However, the aldehyde levels reported here are several orders below those reported for dry puff conditions that reach several hundred µg/g up to 48 mg/g for formaldehyde in some studies21.
Significant aldehyde levels were not generated by the pneumatic atomizer at 140 °C, even though in non-e-cigarette settings formaldehydes can be generated from PG and glycerol at 133 °C, likely because they observed thermal degradation into aldehydes only after 48 h of heating17, and the e-cigarette setup apparently did not substantially accelerate this slow reaction. However, our finding is consistent with a study by Wang et al.22 who only found significant formaldehyde and acetaldehyde levels in the aerosol generated from a PG/glycerol mixture in a chemical reactor when the temperature was equal or above 215 °C. However, in contrast to our study, they did not find acrolein at 215 °C, perhaps because their e-liquid did not contain flavoring chemicals.
The total metal concentrations (9 metals) of the aerosols from the pneumatic atomizer were much lower than those in aerosols from the Mod device operated at 50 W; however, the total metal concentrations, while lower, were comparable to those in e-cigarette aerosols when the device was operated at the lowest power setting of 30 W. The two most abundant neurotoxic metals we found in the aerosol from the e-cigarette we used in this work, Ni, Cu, and Pb, were substantially lower in the aerosols from the pneumatic atomizer than in the e-cigarette aerosols and, in fact, close to or below the MDLs. The levels of Cr and Mn generated by the commercial e-liquid by the pneumatic atomizer, however, were similar to those in the e-cigarette aerosols and were probably introduced by the pneumatic atomizer, because their aerosol levels are higher than their levels in the unused e-liquid. Notably, although we did not perform metal speciation analyses, a recent study reported that Cr is mainly present in its non-toxic form Cr(III), in e-cigarette aerosol23. In contrast, there is no information available regarding Mn speciation or oxidation state in e-cigarette aerosol, while this would be important in determining its potential toxicity.
Concentrations of Cr, Mn, Ni, and Pb were substantially higher in the aerosols generated with the pneumatic atomizer from the commercial e-liquid compared to the unused commercial e-liquid, and Ni levels were amplified the most, about 100 times. This suggests that metals were released from the pneumatic atomizer, even though levels were still lower than in real-world e-cigarette aerosol. The particularly elevated Cr and Ni levels make sense as these elements are frequently used in stainless steel formulations, e.g., in the popular 304 stainless steel, which must contain 18–20% Cr, 8–10.5% Ni, and only 2% Mn24. However, we did not observe elevated Cr and Ni levels when the pneumatic atomizer was fed with the PG/VG mixture, perhaps because flavoring chemicals in the commercial e-liquid could promote metal leaching.
The mechanisms of aerosol generation are totally different in e-cigarettes and the pneumatic atomizer we used to simulate e-cigarette aerosols. E-cigarettes basically boil the e-liquid to form a vapor, which subsequently condenses into an aerosol consisting of inhalable e-liquid droplets; whereas a pneumatic atomizer generates liquid aerosol droplets by a high-velocity air jet, which upon impaction with the bulk, breaks the e-liquid into small aerosol droplets (a process which in the aerosol science literature is called atomization). By heating the pneumatic atomizer, aerosol might also be formed, like in an e-cigarette, by vaporization; however, it is beyond the scope of this paper to elucidate the respective contributions.
It would have been interesting to compare the temperature of the externally heated pneumatic atomizer to the coil temperature of the e-cigarette, as coil temperature can significantly affect e-liquid temperature and aerosol contamination; however, we have only information about the operating power of the e-cigarette. Under normal circumstances, the temperature of the e-liquid cannot exceed its boiling point, and aerosol production can be expected to be significant only if the boiling point is reached. However, if a critical heat flux is exceeded at the surface of the heating coil due to a high operating power, film boiling can occur, and the temperature of the e-liquid can exceed its boiling point25,26. This process can explain the higher aldehyde levels that we observed when the e-cigarette was powered with 50 W compared to 30 W.
While low-cost electrochemical sensors, which are meant to measure gas concentrations, are by no means accurate, they proved useful relative measures in our research to estimate aldehyde levels and refine our experiments. We used them to explore in real time the potential presence of aldehydes in e-cigarette aerosol. However, we confirmed our tentative findings by collecting aerosol samples and analyzing aldehyde contents via LC-MS/MS, which is a strength of the study.
Limitations of our approach for generating metal-reduced but aldehyde-rich aerosols include the following: 1) In these aerosols, not all metals are attenuated equally compared to aerosols from a real e-cigarette. However, levels of the particularly neurotoxic Cu, Ni, and Pb are quite low, and Mn levels were low too. 2) We only examined aerosol from one e-cigarette, 3) We compared the atomizer aerosols to aerosols from a Mod device, a device type not widely used1,27,28,29; however, it allows us, in our parallel animal exposures, to examine the biological effects of the presence of nicotine in aerosols from e-liquids that are identical except for nicotine content. In the US, this can be readily done with refillable Mod devices but not with Pods or disposable devices, because they are typically offered only with zero-nicotine content. 4) We puffed the e-cigarette at a flow rate similar to, as of 2024, the one recommended by the Cooperation Centre for Scientific Research Relative to Tobacco (CORESTA)30; however, it has been suggested that sub-Ohm Mod devices (such as the one we used) are typically used at higher flow rates31,32, presumably due to deeper direct-to-lung inhalation33. Since operating a sub-Ohm Mod device at a lower flow rate promotes aldehyde production and metal release due to less air flow cooling31,34, the aldehyde levels we report might overestimate those experienced by sub-Ohm Mod users. 5) We did not measure background aldehyde levels in the laboratory. Hence, the lowest aldehyde levels that were detected in PG/VG-derived aerosols could be substantially affected by background aldehyde contamination; however, this does not affect our main conclusions as the aldehyde levels we sought to establish were so much higher. 6) We did not determine the size distributions of the aerosols from both the pneumatic atomizer and the e-cigarette, information needed to fully explain inhaled doses of aldehydes of metals. 7) The statistical comparisons between the aerosols and unused e-liquids would have benefited from an increased sample size for each condition examined, something we could not afford due to the high cost of the chemical analyses. However, results are overall consistent, particularly the dependence of chemical concentrations on the temperature at which the aerosols were generated, which makes sense, with aldehyde levels increasing with temperature or e-cigarette operating power.
Disclosures
All authors declare no conflicts of interest.
Acknowledgements
This work was supported by NIH grants R01ES032954 and P30ES009089.
Materials
| Name | Company | Catalog Number | Comments |
|---|---|---|---|
| Agilent 8900 ICPMS | Agilent | https://www.agilent.com/en/product/atomic-spectroscopy/inductively-coupled-plasma-mass-spectrometry-icp-ms/icp-ms-instruments/8900-triple-quadrupole-icp-ms? | |
| Aldehyde/Ketone-DNPH standards | Supelco | ERA-028 | |
| Analyst software | Sciex | version 1.6.2 | |
| C18 column | Kinetex | https://www.phenomenex.com/part?partNo=00D-4475-AN | 1.7 µm |
| Collision-type atomizer | TSI | Model 3076 | |
| Commercial e-liquid | Beyondvape, Baldwin Park, California, US | Pale Whale Vixen’s Kiss | purchased with a 6% nicotine content |
| Electrochemical sensor | Sensirion | SFA30 | |
| Extreme-temperature PTFE tubing | McMaster-Carr | 5239K12 | Extreme-temperature hard plastic tubing for chemicals, semi-clear Teflon® PTFE Plastic without Stripe, 3/16" ID, 1/4" OD, |
| Filtered air supply | TSI | Model 3074B | |
| Heating tape | BriskHeat | TBIH051-020LD | Dual Element, Cloth Insulated Heat Tape, 2 inch x 1/2 inch, 156 Watts |
| High-temperature silicone tubing | Quickun | Pure Silicone Tubing, 1/16" ID, 1/8" OD | |
| Mars 6 | CEM | https://cem.com/mars-6 | |
| Microcontroller board | Arduino | Arduino Uno Rev3 | |
| Mod e-cigarette | Eleaf Electronics Co., Ltd., Shenzhen City, China | iStick Pico Plus, Melo 4S Tank, EC-A 0.3 Ω coil | |
| Multiquant | Sciex | version 3.0.3 | |
| NexION 350S ICPMS | Perkin Elmer | NexION 350S | |
| Nitric acid | Fisher Chemical | Optima | Ultra Trace Elemental Analysis grade |
| Platinum resistance temperature detector | Adafruit | PT1000 | |
| Qtrao 6500 LC-MS/MS System | Sciex | https://sciex.com/products/mass-spectrometers/qtrap-systems/qtrap-6500-system | Tandem mass spectrometer |
| Shimadzu LC | Shimadzu | https://www.shimadzu.com/an/products/liquid-chromatography/index.html |
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