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. Author manuscript; available in PMC: 2021 Jul 30.
Published in final edited form as: Chem Res Toxicol. 2018 Dec 19;32(1):130–138. doi: 10.1021/acs.chemrestox.8b00235

Emissions of free radicals, carbonyls, and nicotine from the NIDA Standardized Research Electronic Cigarette and comparison to similar commercial devices

Zachary T Bitzer ‡,#, Reema Goel ∥,#, Samantha M Reilly ∥, Gurkirat Bhangu ∥, Neil Trushin ∥, Jonathan Foulds ‡, Joshua Muscat ‡, John P Richie Jr ∥,*
PMCID: PMC8323638  NIHMSID: NIHMS1727025  PMID: 30525517

Abstract

E-cigarettes (e-cigs) are a diverse and continuously evolving group of products with four generations currently in the market. The National Institute on Drug Abuse (NIDA) standardized research e-cigarette (SREC) is intended to provide researchers with a consistent e-cig device with known characteristics. Thus, we conducted laboratory-based characterizations of oxidants and nicotine in aerosols produced from SREC and other closed-system, breath-activated, commercially available e-cigs (Blu and Vuse). We hypothesized that oxidant and nicotine production will be significantly affected in all devices by changes in puffing parameters. All e-cigs were machine vaped and the aerosols generated were examined for nicotine, carbonyls, and free-radicals while varying the puff-volumes and puff-durations to reflect typical human usage. The data were normalized on a per puff, per gram aerosol, and per milligram nicotine basis. We found that aerosol production generally increased with increasing puff-duration and puff-volume in all e-cigs tested. Increased puff-duration and puff-volume increased nicotine delivery for Blu and Vuse, but not the SREC. We report, for the first time, reactive free-radicals in aerosols from all closed-system e-cigs tested, albeit at levels lower than cigarette smoke. Formaldehyde, acetaldehyde, acetone, and propionaldehyde were detected in the aerosols of all tested e-cigs. Carbonyl and free radical production is affected by puff-duration and puff volume. Overall, SREC was more efficient at aerosol and nicotine production than both Blu and Vuse. In terms of carbonyl and free radical levels, SREC delivered lower or similar levels to both other devices.

Graphical Abstract

graphic file with name nihms-1727025-f0001.jpg

Introduction

E-cigarettes (e-cigs) heat a liquid mixture (e-liquid) that typically contains a humectant such as propylene glycol (PG) and/or vegetable glycerin (VG), nicotine and/or flavorings to produce an aerosol (E-aerosol) that is inhaled by the user.1,2 The first commercially successful e-cig resembled a conventional cigarette in order to mimic the smoking experience. The rechargeable “cig-a-like” has pre-filled cartridges, and no user options to alter the device power or e-liquid ingredients. The e-cig design has evolved rapidly with four generations of devices in the current market.3 The diversity and ever-changing features of e-cig devices have been an unresolved problem for researchers who look to characterize user behavior, evaluate relative harm and study the toxicological aspects of e-aerosols in an analytical way.4–6 The different structural components, including wicks, batteries, and e-liquids, and design variations from device to device complicate investigations into the potential benefits/harm associated with their use. Some research has been done on e-liquids alone using device-independent aerosol generation systems; however, findings may not be directly applicable to actual human usage conditions and related clinical trials.7 To aid researchers, the National Institute on Drug Abuse (NIDA) has worked with NJOY LLC to develop a standardized research e-cigarette (SREC) for use in clinical studies.8 This device is a breath-activated, closed-system that limits the user to only one voltage and the tobacco flavored e-liquid supplied with the device.

To date, a number of different toxic chemicals have been detected in E-aerosols include carbonyls (acetaldehyde, acetone, acrolein, formaldehyde)9–11, metals (cadmium, lead, nickel, tin, copper)12, nicotine, N′-nitrosonornicotine (NNN)13,14, 4-(Methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) and toluene15. We recently discovered that highly reactive free radicals are also abundant in E-aerosols from 3rd and 4th generation devices.16–19 We also found dose responsive increases in the oxidation of biologically relevant lipids (measured as 8-isoprostane and thiobarbituric acid reactive substances, TBARS) resulting from e-aerosol exposures.17 There is also increasing evidence for e-aerosol-induced oxidative stress, oxidative damage and inflammation in cells lines, animal models and humans.20–24 We found that free radical levels were influenced by device characteristics (e.g., voltage of the power supply, heating element resistance) and e-liquid constituents (e.g., PG, VG, flavor additives).17 While the SREC-aerosol output has been analyzed for some chemicals by NIDA, such as carbonyls and nicotine, there is still a need to fully characterize the e-aerosols for other potentially harmful constituents.

The main objective of this study was to examine SREC-aerosols for oxidants, including free radicals and carbonyls, and make direct comparisons to other popular closed-system devices that are often used in clinical trials. For the comparison, we chose Blu and Vuse, two commercially available breath-activated, closed-system 2nd generation devices rechargeable devices. Blu and Vuse are major e-cig brands based on retail sales, popular e-cig brands among middle and high school students.25,26 Also, both Blu and Vuse e-cig have been used in a number of different laboratory studies and clinical trials, making them both useful to compare to the SREC.27–30 In this study, we examined free radical, nicotine and carbonyl delivery from the SREC, Blu and Vuse under different puffing parameters to capture the likely changes across different user behaviors.

Materials and Methods

E-cigarettes

The SREC battery and tobacco-flavored e-liquid cartomizer tanks (1.48% w:w nicotine) were purchased from NJOY LLC in 2017. All other e-cigarettes, including, Blu (Blu Plus+ rechargeable batteries and “Classic Tobacco” flavored tanks containing 2.4% w:w nicotine) and Vuse (Vuse Solo rechargeable e-cigarette batteries and “Original Tobacco Flavor” tanks containing 4.8% w:w nicotine) were purchased from local retailers (Dauphin and Lebanon Counties, PA, USA) in 2017.

Materials

Acetonitrile (ACN) and hydrochloric acid (12 N HCl) were purchased from Fisher Scientific (Pittsburgh, PA) and used as received. 2,4-Dinitrophenylhydrazine (DNPH) was purchased from BOC Sciences (Shirley, NY) and recrystallized before use to remove water as previously reported.31 Diglyme, phenyl-N-tert-butylnitrone (PBN), 2,2,6,6-tetramethyl-1-piperidinyloxyl (TEMPO), hexane, heptadecane, tertbutylbenzene, and dinitrophenylhydrazones of formaldehyde, acetaldehyde, acrolein, acetone, crotonaldehyde, propionaldehyde, and methyl ethyl ketone (MEK) were all purchased from Sigma-Aldrich (St. Louis, MO) and used as received.

E-aerosol generation

E-aerosols were generated using a single-port smoking machine (Human Puff Profile Cigarette Smoking Machine (CSM-HPP), CH Technologies, NJ). Devices were tested using different puff volumes (75.0 mL and 37.5 mL) and puff durations (2.5 and 5.0 seconds), while keeping inter-puff interval (IPI) (30 sec) and puff number (10) constant. E-aerosols were also generated under the NIDA vaping protocol: 3 sec puff duration, 60 mL puff volume, and 30 sec IPI. The “shape” of each puff resembled that of a square wave. We could not test more “extreme” puff volumes due to limitations of the smoking machine, and puff duration due to cutoffs in the e-cig devices.

E-aerosol production measurements

Cartomizers from each device were weighed before and after each vaping session. The loss in mass was used to calculate the mass of e-aerosol generated.

Free radical measurements

Free radicals in the e-aerosols were captured in an impinger containing 0.05 M nitrone spin trap, phenyl-N-tertbutylnitrone (PBN), in 6 mL hexane as previously reported.17 We have used PBN previously to evaluate the radical content in both cigarette smoke and e-cigarette e-aerosols.16,17,31 After each session, hexane was evaporated under vacuum and the residue was reconstituted in 500 μL tertbutyl benzene. The reconstituted solution was placed into high purity quartz electron paramagnetic resonance (EPR) tubes and deoxygenated by a Schlenk line freeze-pump-thaw technique with argon.32

Electron Paramagnetic Resonance (EPR) measurements

PBN radical adduct derived spectra were measured using a Bruker eScan R spectrometer (Bruker-Biospin, Billerica, MA) operating in X-band. The EPR parameters were: microwave frequency, 9.7 GHz; modulation frequency, 86.0 kHz; microwave power, 6.00 mW; scan range, 60G; modulation amplitude, 2.04 G; sweep time, 5.24 s; time constant, 10.24 ms; and conversion time, 10.24 ms. All measurements were conducted at room temperature (22 ± 1°C). As previously reported, quantitation of the free radicals was done using peak height and the values and were compared against a stable radical standard, TEMPO.31

Carbonyl analysis

Carbonyls were derivatized as described previously using a DNPH solution consisting of 1.0 g of recrystallized DNPH in 150 mL ACN, 50 mL of diglyme, and 360 μL of 12 N HCL.33 The e-aerosols generated from the different devices and puffing profiles were captured in 10 mL of the DNPH solution. 500 uL of pyridine was added to the samples to neutralize the solution to stabilize the acrolein-DNPH derivative. The samples were then stored at 4°C in scintillation vials and analyzed within 3 days of collection. Resulting conjugates were analyzed by high performance liquid chromatography with ultraviolet detection (HPLC-UV) using a binary system consisting of two Waters (Milford, MA, USA) 510 pumps, a Shimadzu (Kyoto, Japan) SPD-10A VP UV-Vis Detector, and a Hitachi (Tokyo, Japan) D-2500 Integrator as previously described.34 In brief, the carbonyls were separated by a C18 column (Bondclone, 10 μm × 300 mm × 3.9 mm; Phenomenex, Torrance, CA, USA) using two mobile phases: (A) 30% ACN, 10% tetrahydrofuran (THF), and 1% isopropanol (IPA) and (B) 65% ACN, 1% THF, and 1% IPA. The elution gradient was: 0 min., 100% A; 8 min., 70% A; 16 min., 60% A; 20 min., 54% A; 22 min., 40% A; 25 min., 100% A; and 31 min. 100% A at a flow rate of 1.5 mL/min. The detection wavelength was 365 nm. All sample injections were 20 μL and injected with a Hewlett Packard (Palo Alto, CA, USA) Series 1050 autosampler. All measurements were carried out at room temperature (22±1°C).

Environmental and experimental blank measurements

Environmental blanks to account for day to day variations in laboratory background air levels of carbonyls were run in tandem with our experiments and subtracted from the carbonyls reported from e-cig vaping. For the environmental blank, DNPH was exposed to atmospheric air for the duration of an e-cig run. Experimental blanks where background levels of free radicals and carbonyls were measured at the higher intensity protocol (3 sec puff duration, 75 mL puff volume, and 30 sec IPI) without an e-cig in line to account for background contamination within the laboratory air, smoking machine, and collection equipment (Table S1). We did not observe any free radicals in the method blank confirming that free radicals were generated by vaping e-cigs only. We observed carbonyl levels in method blanks were the same as environmental blanks, and thus indicating no contamination from any equipment used.

Nicotine analysis

Total nicotine from e-aerosols were trapped onto Cambridge filter pads (CFP). Nicotine was then extracted from the CFP, using 20 ml methanol. Hepatadecane was added as an internal standard. Nicotine was then analyzed by gas-chromatography with flame-ionization detection (GC/FID) using an HP 5890 gas chromatograph with separation on an Agilent CP Wax 52 CB column (30 m × 0.25 mm × 0.25 μm) with helium as the carrier gas at a flow rate of 1.2 mL/min. The injector and detector temperatures were held at 240°C and 280°C, respectively. The initial column temperature was 100°C, held for one minute, then heated to 240°C at 10°C/min, and held for 10 minutes before returning to initial conditions.

Data analysis

All experiments were done at least in triplicate. Values for each device were compared to each other within a puffing regiment using a one-way ANOVA and Tukey’s multiple comparisons test via GraphPad (San Diego, CA). NIDA puffing data is presented in three different ways (per puff, per gram aerosol, and per mg nicotine) in provide a full interpretation of the findings.

Results

E-aerosols produced by changing puff conditions

We examined the effect of differing puff volume and puff duration on e-aerosol production from Blu, SREC, and Vuse devices. When puff volume was varied (37.5 and 75 mL) while maintaining a constant puff duration (2.5 seconds), the SREC consistently produced more e-aerosol than the other devices under both puffing volumes (Figure 1). Blu consistently produced the least e-aerosol under both puffing volumes. While there was a trend of increasing e-aerosol production when puff volume was doubled, this increase was only statistically significant for the SREC. When puff duration was doubled (2.5 to 5.0 seconds) while maintaining a constant volume, the SREC again produced significantly more e-aerosol than the other devices. The longer puff durations resulted in a significant increase in e-aerosol production for both Blu and the SREC but not for Vuse. SREC produced 5–14 mg/puff aerosol under non-intense and intense vaping. Overall, the SREC device produced 275–400% more e-aerosol than Blu and 67–250% more e-aerosol than Vuse while Vuse produced 10–200% more e-aerosol than Blu, depending on the puffing conditions.

Figure 1.

Figure 1.

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Aerosol generated per puff as a result of doubling the puff volume or doubling the puff duration. Different letters indicate significant differences (p<0.05) between brands within the puffing conditions. Error bars indicate standard error.

Puffing parameters effects on aerosolized nicotine

The effects of different puff volumes on nicotine delivery for Blu, SREC, and Vuse devices are summarized in Figure 2. Under the lowest puff volume, the SREC released significantly more nicotine than the other two devices and there was no difference between Blu and Vuse. When the puff volume was doubled, nicotine delivery was significantly higher for Vuse than for the others, followed by the SREC and then Blu. When puff duration was varied, nicotine release was consistently higher in Vuse than in the other devices under both conditions. At the longer puff duration, all three devices released significantly different amounts of nicotine with Vuse being the highest followed by Blu and then the SREC. Doubling the puff duration resulted in significant increases in nicotine release for Blu and Vuse while the SREC’s nicotine release remained constant. Thus, the average nicotine yield for the SREC was 72 μg/puff under the three vaping protocols tested.

Figure 2.

Figure 2.

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Nicotine released per puff as a result of doubling the puff volume or doubling the puff duration. Different letters indicate significant differences (p<0.05) between brands within the puffing conditions. Asterisks (*) indicate significant differences (p<0.05) from the 75 mL, 2.5 s puffing conditions within brands. Error bars indicate standard error.

Puffing parameters on free radical production

We find that free radicals are generated by all the 3 closed-system devices tested. Changing puff volume did not significantly affect free radical output in any of the three devices (Figure 3). At the lower volume, radical delivery from Vuse was significantly higher than for SREC or Blu. At the higher volume, the devices were all significantly different from one another with Vuse > SREC > Blu. A similar pattern was observed when puff duration was doubled, except for Vuse, which had significantly higher radical production with the longer duration. SREC produced 0.03–0.05 nmol/puff free radicals under non-intense and intense vaping.

Figure 3.

Figure 3.

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Free radicals produced per puff as a result of doubling the puff volume or doubling the puff duration. Different letters indicate significant differences (p<0.05) between brands within the puffing conditions. Asterisks (*) indicate significant differences (p<0.05) from the 75 mL, 2.5 s puffing conditions within brands. Error bars indicate standard error.

Puffing parameters on carbonyl production

Four different carbonyls were detected in aerosols from all three e-cig devices, with the highest values being observed for formaldehyde followed by acetone, propionaldehyde and acetaldehyde.

For formaldehyde, no differences were observed between the products at either puff volume, and doubling the volume resulted in significant increases for all of the devices (Figure 4a). While there were no differences in formaldehyde production between devices at the lower puff duration, with the higher duration, SREC produced significantly more formaldehyde than the other devices. The doubling of puff duration caused a significant increase in formaldehyde for SREC only. The SREC produced 0.27–2.29 μg/puff formaldehyde under non-intense and intense vaping.

Figure 4.

Figure 4.

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Carbonyl production by closed system devices as a result of different puffing conditions. Changes in carbonyl production as a result of doubling puff volume or puff duration were measured for (a) formaldehyde, (b) acetaldehyde, (c) acetone, and (d) propionaldehyde. Different letters indicate significant differences (p<0.05) between brands within the puffing conditions. Asterisks (*) indicate significant differences (p<0.05) from the 75 mL, 2.5 s puffing conditions within brands. Error bars indicate standard error.

For acetaldehyde, at the lower puff volume, no significant differences between devices were observed (Figure 4b). At the higher puff volume, the SREC produced significantly more acetaldehyde than Vuse while Blu was not significantly different from the others. Doubling the puff volume resulted in significant increases in acetaldehyde for all devices. When puff duration was doubled, significant increases were observed for Blu and Vuse, but not SREC. At the longer duration, Blu and SREC produced significantly more acetaldehyde than Vuse. Specifically, the SREC produced 0.03–0.24 μg/puff acetaldehyde under non-intense and intense vaping.

For acetone, no significant differences between devices were observed under either puff volume (Figure 4c). The doubling of puff volume resulted in significant increases in all three devices. No significant differences between devices as a result of doubling the puff duration were observed. The SREC produced 0.14–0.60 μg/puff acetone under non-intense and intense vaping.

Propionaldehyde levels were not significantly different between devices under either puff volume (Figure 4d). Doubling the puff volume did result in significant increases of propionaldehyde for all of the devices. When puff duration was increased, Blu produced significantly higher levels of propionaldehyde as compared to the other devices.

Doubling the puff duration also significantly increase propionaldehyde formation for all three devices. SREC produced 0.04–0.67 μg/puff formaldehyde under non-intense and intense vaping.

Other carbonyls, including acrolein, crotonaldehyde, and methyl ethyl ketone, were not detected from the products under any puffing conditions tested.

NIDA method and normalization to aerosol and nicotine content

The preference for different e-cigarette devices often comes down to two factors: their aerosol production (“clouds”) or their efficiency in nicotine delivery. Thus, in an effort to examine delivery based upon these potential user preferences and to compare to past studies, we have expressed and compared free radical and carbonyl production on a per puff, per gram aerosol, and per mg nicotine basis (Figure 5). The puffing parameters for this data used the method developed by NIDA for their initial testing of the SREC (puff volume: 60 mL, puff duration: 3 s, interpuff interval: 30 s, number of puffs: 10–25).8 The SREC produced significantly higher levels of aerosol than the other devices followed by Vuse and then Blu.

Figure 5.

Figure 5.

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Aerosol, nicotine, free radical, and carbonyl production on a per puff, per milligram aerosol, and per milligram nicotine basis using the NIDA puffing method of 60 mL puff volume and 3 second puff duration. Different letters indicate significant differences (p<0.05) between brands and error bars indicate standard error.

Nicotine release was the highest with Vuse followed by SREC and then Blu on a per puff basis. When expressed on a per gram aerosol basis, Vuse was significantly higher than the other devices, which did not differ from one another.

Free radical production did not differ between devices on a per puff basis. However, on a per gram aerosol basis, all of the devices showed significantly different amounts of radicals with Blu being the highest followed by Vuse and then SREC. On a per mg nicotine basis, Blu showed significantly higher levels followed by SREC and then Vuse, which did not differ from one another.

Formaldehyde production on a per puff basis showed significantly different levels between devices with Vuse being the highest followed by Blu and then SREC. The same trend was seen when normalizing the data to aerosol production. When normalized for nicotine release, SREC was significantly lower than the other devices while Blu and Vuse did not differ from each other.

Acetaldehyde levels on a per puff basis were not significantly different. On a per gram aerosol basis, Blu and Vuse were significantly higher than SREC but were not significantly different from each other. On a per mg nicotine basis, Blu was significantly higher than SREC and Vuse, which did not differ from each other.

Acetone levels on a per puff basis and a per gram aerosol basis were lower for SREC than for Blu or Vuse, which do not differ from each other. When expressed on a per mg nicotine basis, Blu was significantly higher than SREC or Vuse, which did not differ from each other.

Propionaldehyde levels did not differ significantly between devices on a per puff basis. On a per gram aerosol basis, Vuse was significantly higher than SREC but not Blu. Blu and SREC were not significantly different. On a per mg nicotine basis, Blu was significantly higher than SREC and Vuse, which did not differ from each other.

Discussion

A reference cigarette has been available since 1960s through University of Kentucky to tobacco research scientists.35 The newly developed e-cig reference product, SREC is now available for clinical research to determine toxicant exposures in e-cig users. It is important to evaluate the chemicals in SREC-aerosols and product features that can impact the toxicant exposures. Previous studies have shown that e-cig design features, such as power, temperature, and the content of nicotine and flavors in e-liquids, can impact user puffing behavior and toxicant exposure.7,36–38 Equally important for toxicant exposures are user puffing behaviors, such as puff duration because energy = power × time. Recent studies have shown that changes in puff duration and puff volume can have a significant impact on the overall aerosol output as well as carbonyl production.39,40 Puffing topography is highly dependent on the product characteristics.41 While the SREC power settings (voltage, wattage, and temperature) and e-liquid composition (nicotine strength and flavor) cannot be altered by the user, they can alter user-defined behaviors, such as puff volume and puff duration. Thus, in this study we have evaluated the impact of these latter factors on delivery of important e-cig constituents, including aerosol production, and delivery of nicotine and oxidants in the SREC device in comparison with other popular commercial rechargeable, closed-system devices (Blu and Vuse).

The testing of the SREC-aerosols by NIDA (NIDA vaping protocol) was done under the following puffing regimen: 60 ml/puff, 3 seconds/puff, 30 seconds inter-puff intervals. We incorporated the NIDA protocol to compare our results, and in general our yields for nicotine and carbonyls are very similar. Recent puffing topography assessments in Blu e-cigs users reported mean puff duration of 2.7±1 sec/puff, puff volume of 56±22 ml/puff, puff count of 33±8 puffs, and inter-puff interval of 17±8 sec.28 Our experimental puffing protocols were designed to encompass a range of puffing conditions observed in users, and also within the range technically feasible in the smoke machine and by the devices. In the future, there is a need to develop more standardized protocols to test a variety of e-cigs, and to include intensive puffing conditions to reflect extreme use.

Few details are available from manufactures about the device power and e-liquid composition for most commercial e-cigs, including Blu and Vuse. However, SREC appears to be a much higher-powered device (9 W) as compared to Blu (4.5 W) and Vuse (~5W) based on manufacturer claims.29,42 Consistent with this higher power, we found that SREC produced significantly more aerosol than either Blu or Vuse. In addition, for SREC, but not Blu or Vuse, we found a large increase in aerosol production when the puff time was doubled from 2.5 to 5.0 sec, even though puff volume remained constant. This is likely due to the coil heating up more quickly and reaching higher maximum temperatures in the more powerful SREC device.

There are inconsistencies in the literature on how to report toxicants in aerosols, with data expressed as per session, per puff, per aerosol production, or per day.14,40,43–46 With the new FDA’s tobacco regulatory strategy to provide smokers “satisfying levels of nicotine” from alternative nicotine delivery systems, it is also important to report toxicants in e-cig aerosols relative to nicotine to account for potential user preferences.47 Due to the wide variations in aerosol and nicotine production observed in the three devices used in this study, we report the results as per puff, per mg nicotine, and per gram aerosol under different puffing profiles. It will allow other researchers and regulatory agencies to compare toxicants across studies and even across tobacco products.

The devices tested in this study differed in the nicotine concentration in the e-liquid, with SREC being the lowest (15 mg/mL) compared to Blu (24 mg/mL) and Vuse (48 mg/ml). We found that nicotine yields ranged from 0.01 to 0.26 mg/puff based on the device and puffing protocol. Both puff volume and puff duration had significant impacts on nicotine yields for Blu and Vuse, but not for SREC. When comparing nicotine output of the devices to one another, SREC was consistently higher than Blu under the 2.5 s puff conditions despite SREC having a lower nicotine content in e-liquid. The nicotine yield from Vuse, with highest nicotine content, was significantly lower than the SREC under the lower puff volume. Thus, the variation in nicotine yields cannot be explained solely by the nicotine content of the e-liquid solutions but must be impacted by other device features such as power and temperature. For the SREC, our results indicate that, even under less intense vaping (37.5 mL, 2.5 seconds), nicotine yields (0.10 ± 0.04 mg/puff) are comparable to those from reference cigarettes under the less intense smoking regimen (ISO 3308:2012) (3R4F − 0.08 mg/puff, 1R6F − 0.09 mg/puff).48,49 As the puff volume and puff duration increase, there is no significant increase in the nicotine for the SREC. This would suggest that despite the lower levels of nicotine found in the e-liquid, the SREC more efficiently and consistently delivers nicotine to the user. For Blu, a minimum puff duration seems to be needed to vaporize nicotine as only the higher puff durations produced nicotine (0.11 ± 0.01 mg/puff) in comparable amounts to a cigarette. Interestingly, for Vuse, higher puff volume resulted in comparable yields of nicotine (0.11 ± 0.02 mg/puff) to cigarettes. With larger puff volumes and longer puff durations, Vuse produced nicotine (0.25 ± 0.02 mg/puff) that would be similar to intense cigarette smoking (Canadian Intense T-115:1999) (3R4F − 0.22 mg/puff, 1R6F − 0.28 mg/puff). However, the relationship between aerosol and nicotine generated is unclear. The difference in nicotine output based on the puffing parameters clearly vary widely between devices, most likely due to their different designs, power, and e-liquids. When looking at nicotine on a per gram aerosol basis, we see many of these trends become amplified as increasing puff volume and puff duration produce slight increases for Blu and Vuse. Taken together, it suggests that the SREC is capable of delivering a consistent level of nicotine despite differences in puffing parameters.

Our results suggest that smokers can achieve similar nicotine levels from e-cigs as are delivered from conventional cigarettes. The initial SREC pharmacokinetic data from NIDA indicate that when people switch from their own e-cigs containing 10–20 mg/mL nicotine to the SREC device, their peak serum concentration (Cmax) is higher and time needed to reach the peak (Tmax) is lower.8 Pharmacokinetic data on cigarette smokers who switch to SREC has yet to be provided.

Much of the carbonyl data also showed increases with increasing puff duration. This most likely arises from the coil increasing to higher temperature with the longer puffs. Increases in carbonyl output as a result of increasing temperatures have been well documented.7,50 Carbonyl production for the SREC appeared to be affected by both increases in puff volume and puff duration. When comparing the devices to one another under the different topography settings on a per puff basis, the SREC appears to produce comparable or slightly higher levels of carbonyls than Blu or Vuse. When normalizing for aerosol or nicotine, the SREC then appears comparable or slightly lower than the other devices tested. As discussed earlier, e-aerosol production and nicotine production increase with puff duration and puff volume although the rate at which they increase varies between devices. These findings suggest that, in terms of carbonyl delivery, participants who use the SREC device are not at increased risk of potential harm compared to other e-cig devices currently used in clinical trials.

We have previously established the analytical methods to generate, sample, and quantify free radicals in e-aerosols in advanced generation e-cig devices.17 Modifying the e-liquid in such systems, we have shown that solvents, flavorants, and temperature affect free radical delivery.17,18 When comparing our present results with this previous data in advanced generation devices, we find that the SREC, Blu and Vuse delivered 4–5 fold lower levels of radical than the more powerful devices.

As puff duration and puff volume increased, so did the radical outputs for all three devices. The increase in radical production with puff duration is expected as the longer time allows for the coil to reach a hotter temperature resulting in more radicals being produced. This temperature-dependent increase in radicals has also been observed in temperature-controlled experiments done by our lab previously.17 Normalizing to aerosol production, the opposite trend is observed because as the puff duration and volume increased the amount of aerosol produced. Normalizing radicals per mg nicotine did not affect the trend for SREC. If nicotine is viewed as the driving force for vaping, then, based on our results, the user would receive the same amount of free radicals with the SREC regardless of their puffing behavior. The radical output of the SREC consistently fell between the other two devices on a per puff basis and was typically lower compared to the others when viewing it on a per aerosol or nicotine basis. Solvent ratio and flavorants likely differ between the three e-liquids and could account for some of the variation in free radical yields observed. Based on our results, even though with similar nicotine yields, the free radical and carbonyls produced by these closed system e-cigs are 5–50-fold lower than from commercial cigarette smoke, which will be important to consider for oxidative stress biomarkers in switching studies.31,51 It is also important to remember that e-cig are fundamentally different nicotine delivery devices compared to cigarettes, and caution must be used to directly compare use and dependence in switching studies.52 The changes in aerosol and nicotine yield as a result of puffing parameters are greater than the changes in free radical production and effectively diminish the effects on radicals. It should be noted that the relationship between aerosol production and free radicals is not fully known and assuming that free radical production positively correlates with aerosol production may not be correct. In conclusion, our results show that the use of most types of e-cig devices will lead to free radical generation and thus inhalation of free radicals. The extent of free radical mediated tissue damage will depend not only on the concentration, but also the radical species and penetrance into deep tissues. Indeed, e-cig exposure studies in cell lines, animal models and humans show evidence of systemic and pulmonary oxidative stress.20–22 Even low levels of repeated exposures to oxidants might increase the risks for cardiovascular and respiratory diseases.3

There were some design features that stood out for SREC compared to the other devices. As mentioned previously, the SREC produced the most aerosol under all conditions. The eliquid in Blu and Vuse cartomizers is completely absorbed into the wicking material, while the SREC is a tank-like cartomizer. In our hands, the SREC cartomizer, in some cases, leaked e-liquid at the contact points. This may be a result of a change in viscosity of the e-liquid as it heats during testing; however, future tests would be needed to confirm this. During our experiments, a buildup of e-liquid and/or condensate from the aerosol in the SREC’s mouthpiece and tubing connected to the smoke machine was observed when 20–50 puffs were taken on the device. We suspect this is a result of the SREC’s higher-powered design and greater aerosol production as similar buildup was not observed with the other two devices. This buildup could affect toxicant measurements when larger number of puffs are taken as toxicants could get trapped by the e-liquid/condensate. Thus, to reduce errors and variability in reporting toxicant deliveries, we report only experiments using low numbers of puffs (10 per session) where no visible condensate was apparent.

With the e-cig market rapidly evolving, studies will also be needed to assess harmfulness, satisfaction and appeal of the SREC among current e-cig users compared to their own e-cig. Their device may be a more advanced mod device capable of generating higher level of toxicants, or the increasingly popular Juul, with sleek design that delivers more nicotine.19 Thus, more careful studies are needed to fully characterize SREC’s components and aerosol chemistry. Nevertheless, SREC offers a standardized e-cig device comparable to other prefilled, breath-activated devices that will provide useful clinical data.

Supplementary Material

Supplementary Material

Funding Sources

This work was supported in part by the National Institute on Drug Abuse of the National Institutes of Health and the Center for Tobacco Products of the U.S. Food and Drug Administration (under Award Number P50-DA-036107). The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health or the Food and Drug Administration.

Abbreviations

ACN

acetonitrile

CFP

Cambridge filter pads

CSM-HPP

Human Puff Profile Cigarette Smoking Machine

DNPH

2,4-Dinitrophenylhydrazine

EPR

electron paramagnetic resonance

GC/FID

gas-chromatography with flame-ionization detection

VG

glycerin

HCL

hydrochloric acid

HPLC-UV

high performance liquid chromatography with ultraviolet detection

IPA

isopropanol

IPI

inter-puff interval

MEK

methyl ethyl ketone

NIDA

National Institute on Drug Abuse

NNK

4-(Methylnitrosamino)-1-(3-pyridyl)-1-butanone

NNN

N′-nitrosonornicotine

PBN

phenyl-N-tert-butylnitrone

PG

propylene glycol

SREC

standardized research e-cigarette

TBARS

thiobarbituric acid reactive substances

TEMPO

2,2,6,6-tetramethyl-1-piperidinyloxyl

THF

tetrahydrofuran

Footnotes

Supporting Information. Environmental and experimental blank measurements showing that the environmental blanks used to account for daily environmental variation do not differ from experimental blanks when just air was passed through the DNPH solution in the impinger.

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