The capture and storage of carbon dioxide (CO2) have considerable potential for mitigating climate change. Adsorption is one of the most popular methods for the storage of CO2. The adsorption of CO2 molecules on the hydroxylated (001) surface of kaolinite was investigated using density-functional theory within the generalized gradient approximation and a supercell approach. The coverage dependence of the adsorption structures and energetics was studied systematically for a wide range of coverage, Θ [from 0.11 to 1.0 monolayers (ML)], and adsorption sites. The CO2 was adsorbed on the two-fold bridge-x (see the text for a definition) and the one-fold top-x sites in the bent, recumbent configuration, and on the three-fold hollow-z, two-fold bridge-z site, and the one-fold top-z sites in the vertical configuration. The surface-adsorbed binding site of CO2 was strongest at the bridge-x site and weakest at the top-z site. The adsorption energy increased with coverage, thus indicating the greater stability of surface adsorption and a tendency to form CO2 islands (clusters) with increasing coverage. The other properties of the CO2/kaolinite (001) system, including the different charge distribution, the lattice relaxation, and the electronic density of states, were also studied and are discussed in detail.

Aversion to the dip-lift stunning system and to the inhalation of 70 and 90% carbon dioxide was assessed in 18 halothane-free (NN) and 14 heterozygous halothane (Nn) slaughter weight pigs using aversion learning techniques and behavioural studies in an experimental slaughterhouse. Pigs were subjected to the treatments individually. When the dip lift system contained atmospheric air, the proportion of pigs that entered the crate voluntarily increased on subsequent days, indicating that pigs habituate to the stunning system. Based on the number of attempted retreats, for the first descent into the well with atmospheric air, Nn pigs were more reactive than NN pigs. On repeating the descent, Nn pigs showed greater habituation to the procedure. When the pit contained (either 70 or 90%) carbon dioxide, the time taken to enter the crate and the incidence of pigs that attempted to retreat increased on subsequent days, indicating aversion to the carbon dioxide concentrations. The aversion was higher when the stunning system contained 90 as opposed to 70% carbon dioxide due possibly to increased irritation of the nasal mucosal membranes and more severe hyperventilation. Conversely, a decrease in the concentration of carbon dioxide increased the time to loss of posture and, therefore, lengthened the perception of the aversive stimulus till the animal lost consciousness. These results suggest that stunning with carbon dioxide is not free from pain or distress. The degree of aversion depends on the carbon dioxide concentration. Therefore, if higher concentrations of carbon dioxide are recommended for rapid induction of anaesthesia, it needs to be assumed that this may be more aversive to pigs.

Controlled atmosphere (gas) stunning (CAS) has the potential to improve the welfare of poultry at slaughter but there is a lack of consensus about which gas mixtures are most humane. The aim of this study was to evaluate the welfare consequences of different gas stunning approaches. Individual broilers were exposed to gas mixtures capable of inducing unconsciousness and euthanasia while their behavioural, cardiac, respiratory and neurophysiological responses were measured simultaneously. The approaches investigated included anoxia (N2 or Ar with < 2% residual O2), hypercapnic anoxia (30% CO2 in Ar, 40% CO2 in N2) and a biphasic method (40% CO2, 30% O2, 30% N2 for 60 s followed by 80% CO2 in air). Evaluation of the welfare implications of each approach centred on the likelihood of them inducing negative states or experiences during the conscious phase. Hypercapnic mixtures were associated with strong respiratory responses, while anoxic mixtures induced vigorous wing flapping. Electroencephalogram analysis using the correlation dimension (a non-linear measure of complexity) suggested that anoxic wing flapping occurred during periods in which a form of consciousness could not be excluded. Hypercapnic hyperoxygenation (biphasic approach) exacerbated respiratory responses but eliminated the possibility of vigorous behavioural responses occurring during a conscious phase. The relative importance of respiratory discomfort versus the potential to induce significant distress due to convulsive wing flapping and associated trauma is a matter for debate. We argue that respiratory discomfort is unpleasant but may be preferable to the risk of vigorous wing flapping and associated injury while conscious in poultry during CAS.

This pilot experiment was conducted to ascertain whether CO2-enriched high expansion foam could be an acceptable and efficient alternative in emergency killing of poultry. This method could have wide-ranging applications but with particular emphasis on small (backyard) flocks, free-range sheds or open (naturally-ventilated) housings. The objectives of the study were as follows: 1) to determine whether the injection of foam and being covered with foam leads to fear or panic reactions in birds; 2) to determine the time taken to render birds unconscious and dead and 3) to determine whether any pathological abnormalities are observed post mortem. Six laying hens were individually exposed to increasing levels of CO2 foam with an expansion rate of 300:1. The test box containing individual birds filled with foam within 30 s. During foaming, two out of six birds tried to escape from the test box (1–2 attempts per bird). Apart from displaying greater alertness, birds showed no aversive reactions to the CO2 foam. Twenty-to-thirty seconds after being covered with foam, five of the six birds demonstrated one or two forcable or convulsive movements. Movement patterns and muscle jerks immediately following this convulsive movement led us to believe that birds lost consciousness at this moment and, within approximately three minutes, all birds had ceased to have a heartbeat. Macroscopic post mortem examination of the birds revealed no abnormalities and microscopic examination showed moderate bronchiolar bleeding and a small amount of alveolar bleeding. After assessing behavioural parameters, measurements of heart rate and pathological data, it is our conclusion that CO2 foam has the potential to be an acceptable method of killing poultry. It is advisable for this method to be examined on a larger scale in order to assess the implications of physiological (EEG and ECG) measurements on welfare.

The distress experienced by animals during the induction of unconsciousness remains one of the most important and yet overlooked aspects of effective methods of anaesthesia and euthanasia. Here we show that considerable differences exist in the aversive responses elicited by 12 common methods of inhalational anaesthesia and euthanasia in laboratory rats and mice. Carbon dioxide, either alone or in combination with oxygen or argon, was found to be highly aversive to both species. The least aversive agents were halothane in rats and enflurane in mice. Exposing these animals to carbon dioxide in any form, either for anaesthesia or for euthanasia, is likely to cause considerable pain and distress and is therefore unacceptable when efficient and more humane alternatives are readily available.

The severity of respiratory distress occurring prior to loss of posture during exposure to: 20, 30, 40, 50, 60, 70, 80 or 90 per cent carbon dioxide in air; 2 or 5 per cent residual oxygen in argon; 30 per cent carbon dioxide in argon with either 2 or 5 per cent residual oxygen; or 40 per cent carbon dioxide in argon with either 2 or 5 per cent residual oxygen, was subjectively determined in pigs from their behaviour. The results indicated that exposure to 2 per cent oxygen in argon (anoxia) induced minimal respiratory distress, 30 per cent carbon dioxide in argon with 2 per cent residual oxygen induced a moderate distress and exposure to all the concentrations of carbon dioxide in air induced severe respiratory distress in the pigs. From the animal welfare point of view, using 2 per cent oxygen in argon (anoxia) appears to be the optimum choice for gas stunning pigs. Secondly, a mixture of 30 per cent carbon dioxide in argon with 2 per cent residual oxygen is preferred to 90 per cent carbon dioxide in air.

High concentrations of carbon dioxide (CO2), used for killing laboratory rodents, are known to be more strongly aversive to rats than sweet food items are attractive. This study investigated whether the maintenance of a high oxygen (O2) concentration, using a gas mixture of 70% CO2 and 30% O2, would reduce aversion to CO2 during a gradual-fill procedure. Eight male Wistar rats, aged 10 months, were housed individually in an apparatus consisting of two cages, one higher than the other and joined by a tube. In a series of trials, subjects entered the lower cage for a reward of 20 sweet food items. The gas was turned on at the moment the rat started eating the reward items and flowed into the lower cage at a fixed rate. There were four treatments: 1) 100% CO2 at 14.5% cage volume min–1; 2) gas mixture at 14.5% min–1; 3) gas mixture at 21.0% min–1, which delivered CO2 at approximately 14.5% min–1 and 4) air, with each subject tested with each treatment four times. Measures of willingness to stay and eat in the lower cage (latency to stop eating, latency to leave and the number of reward items eaten) were much lower in all three gas treatments than in air, indicating that the CO2 and the CO2 + O2 mixture were both more strongly aversive than sweet food items were attractive. Comparing the gas mixture with 100% CO2, the latency to leave and the number of reward items eaten were slightly higher in the CO2 + O2 mixture at 21% min–1 than in CO2 at 14.5% min–1, indicating that the addition of O2 slightly reduced the aversiveness of CO2 in the gradual-fill procedure. This reduction is not enough to warrant recommending the use of CO2 + O2 mixtures for killing rats.

The aversive effects of 90 per cent argon in air, 30 per cent carbon dioxide in air or 90 per cent carbon dioxide in air were investigated in slaughter weight pigs. Aversion was assessed from their reluctance to enter the three gaseous atmospheres to obtain a reward (apples). The pigs did not show any aversion to the inhalation of 90 per cent argon in air. The majority of the pigs did not show aversion to the presence of 30 per cent carbon dioxide in air. By contrast, the inhalation of 90 per cent carbon dioxide was aversive to the majority of the pigs. Fasting them for up to 24h prior to testing did not overcome the pigs ‘ reluctance to enter an atmosphere containing 90 per cent carbon dioxide.

The objective of this study was to evaluate the effects of chamber stocking rate on facets of animal welfare and efficacy during gas euthanasia of young pigs (Sus scrofa domesticus). Crossbred pigs (390 neonatal and 270 weaned) designated for euthanasia at production farms were randomly assigned to group sizes of one, two, four, or six pigs. Gas euthanasia of each piglet group was performed in a Euthanex® AgPro chamber. The chamber air was gradually displaced with CO2 gas over 5 min to establish an in-chamber concentration of approximately 80% CO2. Pigs remained in that atmosphere for an additional dwell period of at least 5 min. Higher stocking rates were associated with higher CO2 concentrations after gradual fill for both age groups. While there was no evidence of an effect of stocking rate on latencies to loss of posture or last movement in neonatal pigs, there was evidence of an effect on all measured efficacy variables in weaned pigs, with grouped pigs faster to succumb than solitary pigs. This finding is consistent with expected consequences of higher CO2 concentration at increased stocking densities. Aversive states and behaviours of focal pigs in the chamber were scored from video. Weaned solitary pigs displayed a high incidence of pacing and may have experienced isolation distress. Escape attempts were absent in neonates and not linearly affected by stocking rate in weaned pigs. Although the risk of hazardous interactions was correlated with group size, this study provided no evidence that isolation during gas euthanasia would benefit animal welfare.

The stability and uniformity of the following gas mixtures: 90% argon; 85% argon and 15% carbon dioxide (CO2); 70% argon and 30% CO2; 98% nitrogen (N2); 92% N2 and 8% CO2; 90% N2 and 10% CO2; 85% N2 and 15% CO2; 80% N2 and 20% CO2; 70% N2 and 30% CO2; and 90% CO2 by volume in atmospheric air were assessed in a commercial dip-lift stunning system when the cradle was either stationary or in motion. The gas mixtures of 90% argon, 85% argon and 15% CO2, 70% argon and 30% CO2, 85% N2 and 15% CO2, 80% N2 and 20% CO2, 70% N2 and 30% CO2 and 90% CO2 by volume in atmospheric air could be sustained in a commercial dip-lift stunning system. The stability of the gas mixtures 92% N2 and 8% CO2, and 90% N2 and 10% CO2 by volume in atmospheric air were lower than in the previous cases. On the other hand, an N2 concentration higher than 94% by volume in atmospheric air could not be sustained in the stunning system. In addition, gas mixtures of argon and CO2 showed a higher stability than gas mixtures of N2 and CO2. The uniformity at different levels inside the pit (defined as the capacity of the gas to maintain its concentration constant at different levels inside the pit) was higher in 90% argon, or argon and CO2 mixtures and N2 and CO2 mixtures than in 90% CO2. This fact ensures that for the whole time the animals are inside the pit, the same conditions are applied, which is not the case for 90% CO2.

Inhalation of concentrations greater than 30% of carbon dioxide (CO2) by volume in atmospheric air causes aversion in pigs. The objective of this study was to assess, using aversion learning techniques and behavioural studies, the aversion to three alternative gas mixtures of nitrogen (N2) and CO2: 70% N2 and 30% CO2 (70N30C), 80% N2 and 20% CO2 (80N20C) and 85% N2 and 15% CO2 (85N15C). The experiment consisted of two trials of three groups of ten pigs each. Pigs were placed individually at the starting point of the test facility and allowed to enter the crate of a dip-lift stunning system during one control session with atmospheric air and three treatment sessions with one of the gas treatments in each group. When the pit contained any of the three gas mixtures, the time taken to cross the raceway and enter the crate did not increase compared to the control session. However, when exposed to the gas mixtures, the majority (85.80%) of pigs performed attempted retreats in the crate, 22.22% exhibited escape attempts, and 7.91% vocalised, without differences between gas mixtures. The percentage of pigs gasping was higher when exposed to 70N30C compared to 80N20C and 85N15C. The results suggest that pigs show signs of aversion to the inhalation of 15 to 30% CO2 in nitrogen atmosphere compared to atmospheric air but the aversion response did not increase in consecutive sessions.

Severely depressed pigs exhibit differences in a number of important parameters that may affect gas euthanasia, including decreased respiration rate and tidal volume. Hence, the objectives of this study were to assess the efficacy and animal welfare implications of gas euthanasia of suckling pigs with varied disease severity (severely depressed [DP] vs other [OT]). A 2 × 2 factorial design was utilised with two gas types (carbon dioxide [CO2]; argon [Ar]) and two flow rates (G = gradual, 35% box volume exchange per min [BVE min−1]; P = prefill + 20% BVE min−1). Sixty-two pigs were enrolled and tested as DP/OT pairs in each gas treatment combination. Pigs identified for euthanasia were assigned a subjective depression score (0 = normal to 3 = severely depressed). Pigs scored 3 and ≤ 1 were categorised as DP and OT, respectively. Significantly lower respiration, rectal temperature, pulse and weight were observed for the DP pigs relative to OT. Pigs were assessed for behavioural indicators of efficacy and welfare. No differences were observed between DP and OT when using P-CO2 or G-CO2. However in P-Ar, DP had greater latency to loss of consciousness relative to OT (212 [± 22] vs 77 [± 22] s), decreased latency to last limb movement (511 [± 72] vs 816 [± 72] s), greater duration of open-mouth breathing (151 [± 21] vs 69 [± 21] s), decreased duration ataxia (101 [± 42] vs 188 [± 42] s) and decreased righting response (27 [± 11] vs 63 [± 11] s). The G-Ar treatment was removed due to ethical concerns associated with prolonged induction. In conclusion, depression score did not affect pig responses to euthanasia with CO2 gas, but did affect responses to Ar. Furthermore, Ar was associated with a prolonged euthanasia process, including frequencies and durations of distress behaviours.

The most widely used stunning method in sheep is electrical. However, in lambs, this method leads to rupture of the blood vessels, provoking ecchymoses. In pigs (Sus scrofa), the use of CO2-stunning systems has increased in popularity due to positive effects on meat quality and animal welfare (movement of animals in groups). The aim here was to assess the effectiveness of a progressive exposure to 90% carbon dioxide (CO2) in inducing unconsciousness in lambs (Ovis aries) through changes in the middle latency auditory-evoked potentials (MLAEP) of the central nervous system (CNS), blood parameters (pH, carbon dioxide partial pressure (pCO2), oxygen partial pressure (pO2), oxygen saturation (SatO2) and bicarbonate (HCO3J, behaviour (head-shaking, sneezing, gasping and gagging) and physiological reflexes (corneal reflex, breathing and sensitivity to pain). Fourteen male lambs of the Ripollesa breed, weighing between 19 and 25 kg were progressively exposed to an atmosphere of 90% CO2 over 66 s. All blood parameters changed between 23 and 43 s after the onset of the immersion. The MLAEP did not decrease significantly until after 48 s exposure to CO2, suggesting an absence of auditory-evoked brain activity. Before that, lambs exhibited head-shaking and sneezing starting at 10.6 (± 0.77) s, and gasping starting at 20.6 (± 1.36) s. After exposure, all animals showed absence of breathing and sensibility to pain, and 36% of them absence of corneal reflex. The pH and pCO2 recovered basal values at 90 and 120 s, respectively, after the end of the exposure. The burst suppression index (BS%) and the A-Line ARX index (AAI) recovered basal levels at 116 and 159 s, respectively. Exposure to CO2 at high concentration induces effective stunning in sheep for a period of 124 s. However, during exposure, the animals exhibited signs of aversion and breathlessness.

The objective of this study was to assess the aversion to exposure of 90% argon, 70% N2/30% CO2 and 85% N2/15% CO2 by volume in atmospheric air in 24 halothane-free slaughter-weight pigs using aversion learning techniques and behavioural studies in an experimental slaughterhouse. Pigs were subjected to the treatments individually during 2 separate trials of 12 animals each. The time of exposure to the gases was 46 and 32 s, respectively. When the pit contained any of the 3 gas mixtures, the time taken to cross the raceway and enter the cradle (TCREC) increased compared with the training sessions (atmospheric air). The incidence of pigs showing retreat and escape attempts and gasps and the number of times that this behaviour was performed was lower in 90% argon than in the gas mixtures with N2 and CO2. On the other hand, the time to loss of posture was lower with 70% N2/30% CO2 than with argon. The second exposure to all gas mixtures was more aversive than the first and the loss of posture also occurred earlier in the second exposure. In conclusion, pigs showed more aversion to gas mixtures with N2 and either 15% or 30% CO2 by volume than 90% argon by volume.

In Sweden, laying hens are killed using the following methods: i) traditional slaughter; ii) on-farm with CO2 in a mobile container combined with a grinder; or iii) with CO2 inside the barn. The number of hens killed using the latter method has increased. During these killings a veterinarian is required to be present and report to the Swedish Board of Agriculture. Data were registered during four commercial killings and extracted from all official veterinary reports at CO2 whole-house killings in 2008-2010. On-farm monitoring showed that temperature decreased greatly and with high variability. The time until birds became unconscious after coming into contact with the gas, based on time until loss of balance, was 3-5 min. Veterinary reports show that 1.5 million laying hens were killed, in 150 separate instances. The most common non-compliance with legislation was failure to notify the regional animal welfare authorities prior to the killings. Six out of 150 killings were defined as animal welfare failures, eg delivery of insufficient CO2 or failure to seal buildings to achieve adequate gas concentration. Eleven were either potentially or completely unacceptable from the perspective of animal welfare. We conclude that, on the whole, the CO2 whole-house gas killing of spent hens was carried out in accordance with the appropriate legislation. Death was achieved reliably. However, there remain several risks to animal welfare and increased knowledge would appear vital in order to limit mistakes related to miscalculations of house volume, improper sealing or premature ventilation turn-off.

The objectives of this research were to evaluate the effect of age and delivering method during carbon dioxide (CO2) euthanasia on the welfare of pigs (Sus scrofa). In Experiment 1, pigs aged 1, 2, 3, 4, 5 and 6 weeks (GRAD; n = 5 pigs per age) were placed in a chamber gradually filled with CO2 released at a flow rate of 20% chamber volume per min. In Experiment 2, three week old pigs were placed in a chamber pre-filled with 100% CO2 (PRE; n = 5). In both experiments, plasma cortisol concentrations were measured before and after exposure to CO2. Behaviours indicative of stress and insensibility were recorded continuously during gas exposure; panting, open-mouth breathing, righting response, escape attempts, loss of posture, muscular excitation and respiratory arrest. Cortisol concentrations were elevated in pigs after CO2 euthanasia, regardless of age or gas delivery method. The behavioural response to CO2 was not affected by pig age. Latency to display panting, righting response, escape attempts, loss of posture, muscular excitation and respiratory arrest were shorter for PRE than GRAD pigs, but, duration of escape attempts and the cortisol response to euthanasia were similar between PRE and GRAD pigs. However, pigs placed in a chamber pre-filled with CO2 spent a greater proportion of time prior to loss of posture displaying escape attempts. Regardless of age or induction method, exposure to CO2 causes behavioural changes indicative of stress prior to loss of consciousness, therefore there is a need to continue to evaluate alternative methods of euthanasia.

Exposure to high concentrations of CO2 is a common means of stunning and killing laboratory rodents. However, there is concern regarding the potential for animals to have aversive experiences, such as pain or breathlessness, prior to loss of awareness. This preliminary study evaluated the electroencephalographic (EEG) responses of rats (Rattus norvegicus) to CO2 inhalation, using a method based on a minimal anaesthesia model previously used to assess nociception in mammals. Fifteen adult female Sprague-Dawley rats were anaesthetised with halothane in oxygen and maintained at a minimal plane of anaesthesia. EEG was continuously recorded throughout a 10-min baseline period followed by sequential exposure to 5, 15, 30 and 50% CO2. The EEG summary variables median frequency (F50), 95% spectral edge frequency (F95) and total power (PTOT) were derived from the raw EEG. The F50 of the EEG, a sensitive indicator of nociception, increased significantly above baseline during exposure to 15% CO2, suggesting this concentration was noxious to rats. This is consistent with behavioural aversion in rats at around the same CO2 concentration. Stimulation of the rat mucosal nociceptors only occurs at CO2 concentrations of 37% or greater; therefore, it is hypothesised that the observed response was a result of what would have been CO2-induced aversive respiratory sensation in conscious animals, rather than pain. This study provides some evidence that an anaesthesia model may be utilised to study the nocuous effects of low-moderate CO2 exposure in rodents.