Apparatus at IU for active, time-resolved measurements of gas concentrations with circular flow

At IU, we constructed an experimental apparatus to assess the loss of CH₄ in an active (i.e., with pumping of air) and time-resolved manner with or without added radiation from radon isotopes and their progeny (Fig 1). The use of pumping qualifies this method as active and time-resolved in contrast to passive measurements of radon that integrate over time [19]. Approximately 6 L of air was recirculated in the sealed apparatus that included (i) a glass tube with optional thorium carbonate to generate ²²⁰Rn (also called thoron), and (ii) a glass tube containing uranium ore to generate ²²²Rn, with an overlying layer of coconut charcoal to limit the escape of co-produced, short-lived ²²⁰Rn. Blank experiments without elevated radiation identified a reproducible loss of CH₄ (likely by diffusion through polymer tubing within the sealed analytical SARAD RTM 2200 instrument) that was subtracted from all other experiments at IU to arrive at net CH₄ losses that are due to other factors, such as radiolysis or microbial methanotrophy.

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Fig 1. Experimental setup for time-resolved measurements of gas concentrations with circular flow at Indiana University.

Approximately 6 L of air was recirculated in a sealed apparatus to assess the loss of CH₄ with or without added radiation from radon isotopes and their radioactive progeny. At the beginning of each experiment, the trapped air was slightly enriched in CH₄ (and CO₂, except for experiments with soils), followed by hourly measurements of gas concentrations over a few days to weeks. Radon ²²²Rn was generated by uranium ore while charcoal retained ²²⁰Rn (a). The air intake of the 3-neck 5-L glass flask was directed to the bottom of the flask with a plastic insert to facilitate the mixing of air (b); thorium carbonate is not shown. Depicted components of the apparatus (c) are not drawn to scale.

https://doi.org/10.1371/journal.pone.0206506.g001

We conducted a number of experiments at IU to assess the importance of α-radiation intensity, relative humidity, and the presence or absence of soil on CH₄ dynamics. Moisture is critical for the emanation efficiency of radon isotopes from solid sources (i.e., the escape of noble gas radon atoms from the interior of minerals into H₂O-containing pore space via recoil subsequent to radioactive decay of parental nuclides; e.g., [20, 21]) and for stabilizing ions and radicals in air. Individual experiments differed in terms of their optional use of elevated humidity, thorium carbonate, and gas flowing through the tube containing uranium ore. The trapped ~6-L volume of air was initially spiked with CH₄ from natural gas to ~70 ppmv and with CO₂ to ~5,000 ppmv (except for experiments with soils) to distinguish it from room air and to increase the analytical precision during the time-series of measurements that lasted over a few days to weeks. Elevated CO₂ concentrations are typical for many cave environments [10].

Most experiments at IU discriminated between α-radiation from radon ²²²Rn versus thoron ²²⁰Rn. Whereas radon ²²²Rn with a half-life of 3.83 days is relatively homogeneously distributed in cave air (also in our apparatus), the much shorter lived thoron ²²⁰Rn with a half-life of only 55.6 s [22] cannot travel far from its parent nuclei residing in minerals [19], thus thoron’s highest concentrations in cave air are near cave walls and the floor. The higher α-decay energy of ²²⁰Rn (6.3 MeV) relative to ²²²Rn (5.49 MeV) prompted us to design experiments for separate examinations of the ability of both radon isotopes to trigger the oxidation of CH₄. The more energetic α-decay of thoron ²²⁰Rn should ionize air more efficiently than ²²²Rn. Thoron was generated from thorium carbonate that was optionally loaded into a glass tube attached to the round-bottom flask. In other experiments, ²²²Rn decay measuring up to 327 kBq m^-3 was produced in-situ in the glass apparatus by uranium ore chips (Fig 1). Escape of co-produced thoron from ore was reduced by using a layer of coconut charcoal in the upper part of the glass tube as a filter [23]. The resulting adsorption of ²²⁰Rn on charcoal increased the residence time in the glass tube and let ²²⁰Rn decay before it could enter the 5-L glass flask.

We quantified the concentrations of ²²²Rn, ²²⁰Rn, CH₄ and CO₂ during experiments at IU at an air flow rate of ~0.2 L min^-1 once every hour while operating the diffusion pump in the SARAD RTM 2200. ²²⁰Rn radiation intensity was either measured via α-spectroscopy at a faster flow rate of 1 L min^-1 in 10-min increments (n ≥ 10) while temporarily operating the more powerful membrane pump, or values from flow rates ≤0.2 L min^-1 with the diffusion pump were doubled to adjust for fast ²²⁰Rn decay (see S1 File for detailed control experiments and graphed data). Elevated relative humidity fosters the stabilization of ions in air via attachment to clusters of water molecules and may enhance the ability of ions to trigger oxidative degradation of CH₄ (discussed in [10]). Therefore, at IU we recorded humidity in the apparatus along with temperature, air pressure, flow, and battery voltage on an hourly basis. The accuracy of data from the SARAD RTM 2200 was independently evaluated via direct comparison with a newly manufactured and factory-calibrated Thoron Scout instrument (SARAD GmbH, Dresden, Germany; details available in S1 File).

We conducted a number of experiments at IU to test for the effects of radiation and microbial activity on CH₄ dynamics in our experimental apparatus. Multi-day time-series of data were collected in closed-circuit air reflux mode (i) as duplicated blank experiments without added radon or thoron, (ii) with enhanced ²²⁰Rn concentration in dry or moist air, (iii) with enhanced ²²²Rn concentration in dry or moist air, and (iv) with jointly enhanced ²²⁰Rn and ²²²Rn concentrations in moist air to depict an extreme scenario where cave air had a highly elevated α-radiation level. Furthermore, (v) we tested for CH₄ oxidation after placing moist soils, which we assumed contained methanotrophic bacteria (MOB), into the 5-L glass flask, without elevated radioactivity. Certain impurities in industrially conditioned natural gas may act as MOB inhibitors, for example acetylene and carbon monoxide (p. 335 in [24]). As a precaution, the CH₄ spikes in experiments employing two different soils were derived from gas that was collected from a natural seepage of shale gas in New York State [25]. Natural shale gas is not known to contain acetylene or carbon monoxide.

Gas-tight terrarium experiments at RHUL

Experiments at RHUL at atmospheric CH₄ abundance used a gas-tight glass terrarium (i.e., an aquarium without water holding a volume of 13.45 L; Fig 2) with a hermetically sealing glass lid. Two air-tight gas ports allowed the withdrawal of 1-L air samples into Tedlar bags without changes in atmospheric pressure. Fragments of uraninite-bearing pitchblende served as a source of radioactivity. An AlphaLab Air Ion Counter with an integrated fan was placed in the terrarium to measure the abundance of ions in air in 30-s intervals. The α-radiation was quantified on 1-h intervals with a Canary Pro monitor (Airthings, Oslo, Norway) via α-spectrometry. Gas samples in Tedlar bags were analyzed for CH₄ mole fractions with a Picarro G1301 CRDS (Cavity Ring-Down Spectrometer, Picarro Inc., Santa Clara, California, USA).

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Fig 2. Gas-tight terrarium experiments at Royal Holloway University of London.

(a) A hermetically sealed glass terrarium was filled with laboratory air containing atmospheric CH₄. A radon monitor provided data on ²²²Rn abundance, while an AlphaLab Air Ion Counter measured the concentration of negative ions. After ~6 h into the RHUL experiment #1, the placement of a beaker filled with deionized, warm water elevated the relative humidity to ˃ 85%. At the same time, two fragments of pitchblende (containing uraninite as a radiation source) were placed into the terrarium to generate ²²²Rn. Tedlar bags in the terrarium are not shown in the photograph. (b) Diagram of the sampling procedure to collect <1-L aliquots of air from the terrarium in RHUL experiment #2. This experiment lasted for 76 h and 50 min and reached a ²²²Rn-based radiation level in excess of 50 kBq m^-3 after 5 h.

https://doi.org/10.1371/journal.pone.0206506.g002

The initial RHUL experiment #1 (Fig 2A) assessed the production of negative ions and the abundance of ²²²Rn over ~6 h (i.e., stage 1) without either pitchblende or a beaker with water in the terrarium that had been flushed initially with laboratory air, and subsequently for ~15 h in the presence of pitchblende and a beaker with 130 mL of 38°C warm water in the terrarium (stage 2).

The subsequent RHUL experiment #2 (Fig 2B) in the same terrarium included monitoring of the CH₄ mole fraction of laboratory air sealed in the terrarium where pitchblende and a beaker with 130 mL of water (initially at 38°C) had been placed to provide for elevated radioactivity and relative humidity. Elevated relative humidity was needed to simulate cave conditions. The AlphaLab Air Ion Counter failed to provide useful data due to static interference with Tedlar bags. The second experiment lasted for 76 h and 50 min and reached a ²²²Rn-based radiation level in excess of 50 kBq m^-3 after 5 h. Approximate 1-L aliquots of air sampled from the terrarium were analytically compared with aliquots of exterior laboratory air on four occasions.

Active time-series measurements with circular flow at IU

Our controlled experiments with and without ²²⁰Rn and/or ²²²Rn were designed to directly test whether or not radiation can oxidize CH₄ in cave air on ecologically relevant time scales (i.e., hours to days). We relied on comparisons of CH₄ inventories in experiments with (i) high radiation intensity from in-situ generated ²²⁰Rn and/or ²²²Rn with those from (ii) duplicate blank experiments with no artificially enhanced radiation to demonstrate the sensitivity of our setup to detect CH₄-losses. In addition, we conducted (iii) two experiments with moist soils in the absence of added radon isotopes to assess the potential for environmental microorganisms (i.e., MOB) to remove CH₄ as has been demonstrated elsewhere by members of our research team [14, 18]. The comparisons among experiments covered a common range of CH₄ concentration and thus only differed in the lengths of their time windows needed to lower the CH₄ concentration from the upper to the lower threshold (i.e. yellow rectangle in Fig 3A). The ‘common window’ of CH₄ decline for all 11 experiments maximized the data available for comparison.

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Fig 3. Time-series experiments at Indiana University.

Results from time-series experiments to test for abiotic (radiation) and biotic (CH₄ oxidizing bacteria) factors on CH₄ dynamics. (a) The decline of CH₄ concentrations in experiments #1 to #9 (without soil) followed similar trajectories depending on original concentrations, despite major differences in radiation intensity (see the yellow rectangle that identifies a window of CH₄ concentrations that is common to all experiments). Declining CH₄ concentrations are independent of the intensity of α-radiation. Blank experiments #1 to #4 without elevated radiation identify a reproducible loss of CH₄ by diffusion that was subtracted from all other experiments to arrive at net losses that are due to other factors, such as radiolysis or microbial methanotrophy. (b) The decline of CO₂ concentrations in a range of experiments without soil followed similar patterns. In addition to loss due to diffusion through plastic, it was likely influenced by adsorption, solution in water, or possible chemical uptake. In addition, CO₂ was generated from moist soils in experiments #10 and #11. (c) Radon ²²²Rn and (d) thoron ²²⁰Rn concentrations partially depended on relative humidity; soil no. 2 in experiment #11 generated low levels of ²²²Rn over time presumably due to traces of uranium in minerals; ²²⁰Rn concentrations are original data from low flow rates at <0.2 L min^-1 when laminar flow conditions in the 5-L glass flask caused heterogeneity and occasional spikes. (e) Noise in relative humidity data partially derived from the automatic battery recharge cycle that influenced the internal temperature of the SARAD RTM 2200 and the algorithm to calculate humidity. (f) Experiments #10 and #11 with moist soils without added ²²²Rn or ²²⁰Rn resulted in a long-term exponential decline of CH₄ concentrations while CO₂ was generated biologically.

https://doi.org/10.1371/journal.pone.0206506.g003

Multiple trials in our experimental apparatus revealed that CH₄ dynamics were unaffected by radiation within the precision of measurements. Repeat blank experiments with dry (experiments #1 and #2) or moist air (experiments #3 and #4) without artificially elevated radon or thoron concentrations resulted in reproducible and systematic small losses of both CH₄ and CO₂ over time (Fig 3A and 3B; Table 1A). Although radon isotopes, CH₄ and CO₂ could not diffuse through glass and metal in our apparatus, the SARAD RTM 2200 and its Axetris laser OEM Module LGC F200 methane detector were internally and externally connected to glass and metal components with short segments of various types of clear polymer tubing (Fig 1) that resulted in slow losses via gas diffusion through polymers. The rate of diffusion across a layer of polymer is dependent on the difference in partial pressures between the interior and exterior air, and hence the rates of CH₄ and CO₂ losses via diffusion over time follow curves that asymptotically approach equilibria (Fig 3F). At a CH₄ concentration of ~59 ppmv in the apparatus (i.e., the midpoint of the common CH₄ range; Fig 3A) and outside air with ~1.85 ppmv, the mean CH₄ diffusive loss from air in the apparatus during blank experiments #1 to #4 consistently amounted to 0.39 ppmv h^-1 regardless of humidity and small variations in room temperature and air pressure (Table 1A; S1 File). Such a loss of gas over time could theoretically result from a small internal leak in the system. However, the non-parallel pattern of CO₂ losses in blank experiments (Fig 3B) is inconsistent with a leak and instead argues for varying diffusivity of the polar molecule CO₂ through permeable material at different humidities. The observed degree of CH₄ loss from the system was unavoidable and had to be subtracted from the observed bulk CH₄ losses in experiments with enhanced radiation and soils to arrive at any specific losses that are due to radioactivity or presumed microbial methanotrophy.

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Table 1. Overview on individual experiments performed at Indiana University to constrain the consumption of methane over time.

https://doi.org/10.1371/journal.pone.0206506.t001

There was a comparable loss of CH₄ in recirculating air for all experiments without soil, regardless of the absence or presence of radiation from ²²⁰Rn, ²²²Rn, or both ²²⁰Rn and ²²²Rn, in dry or moist air (Fig 3A). The time needed to cross the ‘common window’ of CH₄ decline from 67.2 to 50.9 ppmv was not shorter when radiation from ²²⁰Rn and/or ²²²Rn was added (Table 1A, 1B). The slopes of lines representing CH₄ decline within the common window in Fig 3A were not higher for experiments with elevated radiation (mean ~0.38 ppmv h^-1) than for blank experiments without added radon isotopes (mean ~0.39 ppmv h^-1; Table 1A and 1B). The mean levels of added radiation from ²²⁰Rn, and especially the cumulative radiation in experiment #9 from simultaneously added ²²⁰Rn and ²²²Rn, ranged between ~50 and 535 kBq m^-3 after doubling of experimental ²²⁰Rn values that were measured at ≤0.2 L min^-1 (Table 1B) and thus always exceeded the radiation levels reported in cave air [26], including the air in all Spanish caves where abiotically driven CH₄ oxidation due to radiolysis has been reported [10]. For example, the average rate of CH₄ consumption in Spanish Altamira Cave air of -0.03 ppmv h^-1 occurred at a maximum ²²²Rn radiation level of ~6 kBq m^-3, which is roughly one to three orders of magnitude less than the radiation in any of our experiments with added radon isotopes (#5 through #9). Thus, in terms of radiation intensity, our experiments represent an extreme test of the radiolysis hypothesis. Only the air in shafts of underground uranium mines has been observed to reach even higher radiation levels of one million or more Bq m^-3 [27].

The consistent pattern of CH₄ decline in our experiments without soils can be better appreciated in light of the observed CO₂ dynamics (Fig 3B). CO₂ is more polar than CH₄, can be more easily adsorbed on surfaces, and is more water-soluble and reactive than CH₄. Therefore, it is possible that changes in room temperature (21.1 to 27.5°C) and atmospheric pressure (96.7 to 99.3 kPa) may have affected adsorption and solubility of CO₂ during our experiments. Moreover, after one week of measurements with a wet paper tissue in the 4-L glass flask without soil, fungi had discolored the paper tissue and metabolically generated CO₂, thus partially stabilizing the CO₂ partial pressure (experiment #6, Fig 3B), apparently without affecting the CH₄ decline (Fig 3A). The paper tissue had been hung by a thread from the central glass stopcock to maximize surface area and to avoid any anoxic microenvironments that could facilitate biological methanogenesis (Fig 1). Subsequent experiments in moist air without soil replaced the wet paper tissue with added deionized water at the bottom of the 4-L glass flask. Experiments with soils initially generated CO₂ via microbial and fungal remineralization of soil organic matter, followed after several days by a decline due to diffusive loss of CO₂.

In the two experiments with moist soils, we documented a CH₄ loss of of ~0.09 ppmv h^-1 within the common window of CH₄ concentration decline (Fig 3A), as determined by subtracting the diffusive CH₄ loss in blank experiments from the bulk CH₄ loss in experiments #10 and #11 with soils (Table 1). It is well established that heterogeneously distributed methanotrophic biofilms in the subsurface [28] are capable of scavenging CH₄ from the atmosphere (e.g., [29, 30]). Soil gas can often reach ²²²Rn radiation levels of many thousand Bq m^-3, depending on local geology [31, 32]. If radiolysis would indeed be able to trigger fast oxidative decay of CH₄ in soil gas, such an important CH₄ sink in dry soils without abundant methanotrophic activity would likely have been documented. Also, radiolysis would compete with methanotrophs in moist soils for CH₄ and would have been identified as a factor in soil CH₄ studies.

Experiments in gas-tight terrarium at RHUL

The first stage of experiment #1 at RHUL (Figs 2A and 4) established background conditions for the abundance of negative ions (~3800 ions cm^-3) and the concentration of ²²²Rn (17 to 51 Bq m^-3) in laboratory air at temperatures from 21.4 to 21.7°C and relative humidities from 26.6 to 29.0%. After the onset of stage 2, the placement of pitchblende and a beaker with 130 mL, 38°C warm water into the sealed terrarium strongly increased the abundance of negative ions in air (up to ~200,000 ions cm^-3) and the concentration of ²²²Rn (~118 kBq m^-3). The relative humidity exceeded 85%, and the air temperature intermittently rose by 5°C. The measurement uncertainty of the Canary Pro radon monitor increased with the ²²²Rn radiation level (Fig 4). However, the factory-documented uncertainty at the highest measured radiation level and the steadily increasing abundance of negative ions in air suggested that after a run time of ~17 h, the ²²²Rn-based radiation level exceeded 100 kBq m^-3 (Fig 4; data shown in S1 File).

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Fig 4. Experiment #1 at Royal Holloway University of London.

The experiment proceeded in two stages where the first stage established background conditions in a hermetically closed terrarium at low relative humidity and in the absence of artificially elevated concentrations of ²²²Rn. In the second stage, the addition of pitchblende and a beaker with hot water provided a source of ²²²Rn and high humidity to simulate conditions characteristic of cave environments. Although the final ²²²Rn concentration exceeded 100 kBq m^-3 and was thus higher than in most caves, the elevated radiation and ionization of air in the terrarium was unable to lower the atmospheric abundance of CH₄ over 77 h in the subsequent RHUL experiment #2 (Table 2).

https://doi.org/10.1371/journal.pone.0206506.g004

Experiment #2 at RHUL (Fig 2B) used the same sealed terrarium with pitchblende and high humidity to monitor and compare the CH₄ mole fractions in the air of both the terrarium and the outside laboratory air over ~77 h. The Canary Pro radon monitor in the terrarium indicated an increase in ²²²Rn over time parallel to RHUL experiment #1. After 5 h into RHUL experiment #2, the ²²²Rn-based radiation in the terrarium was consistently > 50 kBq m^-3. Despite high levels of ionization and ²²²Rn-based radiation in the terrarium, the CH₄ mole fraction of 1.9941 ± 0.0036 ppm in terrarium air after being sealed for ~77 h was indistinguishable from the starting value of 1.9971 ± 0.0122 ppm within the uncertainty of measurements (Table 2).

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Table 2. Results of experiment #2 at RHUL to assess the consumption of CH₄ over time in the presence of elevated ²²²Rn concentrations and high relative humidity.

https://doi.org/10.1371/journal.pone.0206506.t002

Synopsis of combined results

The absence of any experimental evidence for accelerated loss of CH₄ in the presence of elevated radiation makes it highly unlikely that radiation from radon isotopes is important in nature where ²²⁰Rn and ²²²Rn concentrations are typically much lower. Our data indicate that natural radiation in cave air cannot be responsible for the rapid consumption of CH₄ in air on time-scales of days, even in caves with high relative humidity. The same conclusion had been reached from earlier laboratory experiments [18] and from observations in Australian cave air [15].

Many caves experience seasonally different degrees of venting and even reversals of air flow, which results in differences in air temperature and humidity and is difficult to simulate in laboratory experiments. Still, most cave environments at sufficient distances from cave entrances and vent holes are thermally buffered by surrounding rock and therefore do not express the relatively high diurnal and seasonal temperature and humidity variations as outside environments. Our experiments in laboratories were conducted at relatively constant room temperatures similar to many cave environments. Room temperatures in air-conditioned laboratory buildings are similar to actual temperatures in sub-tropical and tropical caves [14]. The use of water and moist soil in many of our experiments simulated the range of humidity in natural cave air. One possible caveat in terms of dissimilarity between our laboratory settings and actual caves may be the fact that our experiments allowed daylight to reach our experimental setups. However, the amount and timing of indirect light (no direct sunshine) was insufficient to let any photoautotrophs (algae) observably grow in our experiments. A necessary difference between air in our experiments at IU and actual cave air was the presence of traces of CH₄ in our experiments. Some CH₄ was needed to test for possible radiolytic destruction of CH₄. In contrast, most natural cave air is depleted in CH₄ relative to outside air. We conclude that the experimental conditions during experiments at IU and RHUL were reasonable approximations to simulate cave conditions. In the open atmosphere, solar radiation is mainly responsible for the generation of OH• radicals ([6], and refs. therein) that are the longest-lived potential radical reactant with CH₄ in air. Subterranean radiolysis by radioactivity involves far more energy than photochemical dissociation of molecules by solar radiation, hence the speciation of resulting ions and radicals is different. A host of highly energetic, short-lived ions and radicals other than OH• is generated in subterranean air. The first abstraction of an H atom from CH₄ requires a far higher activation energy than those of H atoms from methyl CH₃ and methylene CH₂ moieties. We argue that cave environments with elevated radioactivity may host short-lived, yet highly energetic radicals and ions that can supply the needed activation energy for first H-abstraction from CH₄ more efficiently than OH• in the open atmosphere. Thus, the application of kinetic and energetic findings of photochemical CH₄ oxidation in the open atmosphere may not be warranted for subterranean environments.

The α-radiation level in cave air is typically higher than in the open atmosphere because cave air is relatively close to rock and sediment surfaces with minerals harboring radioactive nuclides. The ionization rate in air via ²²²Rn radon decay is larger close to the ground, as reported for a Finnish forest [33], a spa [34], and in houses [35]. The effect is due to (i) strongly elevated radon concentrations in the air in porous, uranium-containing substrates and the rapid dilution of radon above surfaces upon mixing with the open atmosphere, especially during windy conditions. In contrast, cave air far from cave entrances is typically less turbulent and allows for a more even distribution of ²²²Rn in cave air. (ii) Short-lived ²²⁰Rn will always exhibit a greater abundance in air close to its parent nuclides in soil, rock, cave walls and floors [19]. Regardless, even exceptionally high combined radiation levels of ²²⁰Rn and ²²²Rn provided no evidence for accelerated CH₄ oxidation in our experiments.

A plausible reason for slow radiolytic reaction kinetics is the mismatch between the large number of CH₄ molecules in 1 m³ of atmosphere containing 1.85 ppmv CH₄ at standard conditions (i.e., ~4.55 · 10¹⁹ molecules CH₄) relative to the small number of radon-related nuclear decay events in the same volume of air (e.g., 10 kBq m^-3 from ²²²Rn resulting from the decay of 10,000 atoms of ²²²Rn per second). The following simplistic numerical example illustrates the lack of feasibility of radiation-induced rapid oxidation of CH₄. If we assume that 1 m³ of atmosphere entering a cave with 10 kBq m^-3, even if every decay of ²²²Rn leads to the oxidation of one molecule CH₄, it would require a geologic time period of ~144 million years to oxidize all CH₄. In reality, the nuclide-specific radiation from the decay of ²²²Rn alone is dwarfed by the total radiation from radon, thoron, their radioactive progeny, and any other radioactive nuclides present in a given environment [19]. S1 File offers alternative calculations based on the assumptions that either (i) all energy from α-decay is exclusively invested in radiolytic dissociation of CH₄ and results in the oxidation of multiple molecules of CH₄ per decay event, or (ii) that only a fraction of the energy from α-decay is dissociating CH₄ in the overwhelming presence of other molecules and atoms. The calculated time periods needed to degrade 1.85 ppmv CH₄ at a ²²²Rn radiation level of 10 kBq m^-3 range from 45.1 to 153,000 years, respectively. Even the most optimistic assumptions cannot speed up the radiolytic reaction kinetics to consume atmospheric CH₄ within hours to days.

We can use the most optimistic scenario for consumption of 1.85 ppmv CH₄ during 45.1 years at 10 kBq m^-3 and calculate a radiation level of ~165 MBq m^-3 that would be required to perform the same task in 24 h, which would be commensurate with kinetic CH₄ observations in caves. Natural radiation levels of a few MBq m^-3 have been measured in air where ²²²Rn emanates through geologic faults from underlying uranium minerals [36]. Radiation levels in the range of MBq m^-3 have been observed in the air of uranium mines [27]. Still, no location is known to offer values close to the required ~165 MBq m^-3. We conclude that there is no natural cave environment on earth where the α-radiation level is strong enough to rapidly degrade CH₄. The same conclusion was recently described in a study that included arguments based on radiolytic kinetics of ion-induced reactions [15] that complement our calculations using α-decay and activation energy.

Subterranean radiation does not provide a mechanism for a fast-acting sink of atmospheric CH₄ that would extend to arid and hyperarid environments, unlike microbial methanotrophy. Our study does not invalidate the geochemical data from previous studies documenting CH₄ dynamics in subterranean ecosystems [10]. We do not call into question the fundamental importance of radiolysis of H₂O (and other air components) and subsequent redox reactions that are documented in the geologic record (e.g., [37]) or the long-term subterranean radiolytic impact on sedimentary organic matter [1]. However, the exceedingly slow chemical rates of reaction caused by natural rates of radiolysis would likely take years to geologic time periods in cave environments to deplete trace amounts of atmospheric CH₄ in cave air. As long as no alternative mechanisms have been identified, microbial methanotrophy serves as the only known fast-acting sink for subterranean CH₄ in the critical and vadose zones.