Pre slaughter factors in pigs on initial pH and temperature: a case study

ABSTRACT: This study evaluated factors associated with the pre-slaughter handling of pigs in a commercial slaughterhouse located in Minas Gerais and evaluated the influence of these factors on the initial pH and temperature of the carcasses. In all, 4449 carcasses were evaluated. The factors evaluated were transport distance, transport duration, loading density, fasting duration, resting duration, average weight of the animals, average temperature in the resting area, and maximum temperature in the resting area. The effects of these factors on the initial (i.e., at 45 minutes after slaughter) temperature and pH (pH45) were measured. The pH45 values were used to determine the occurrence of PSE; the carcasses were classified as PSE when they presented a pH45 < 5.90. Results showed that animals fasted for more than 24 hours were more likely to result in PSE meat (1.41 times more likely), as did animals that were exposed to an average temperature above 24 °C (1.52 times more likely). Except for the average weight of the animals, which did not affect the initial temperature values, all other pre-slaughter factors have influenced both pH45 and the initial temperature values, and the difference of fasting duration and the temperature in the resting area increase the incidence of PSE meat.


INTRODUCTION
Pork is the most consumed animal protein worldwide, and Brazil ranks fourth in pork production and exports.The quality of pork has multifactorial aspects, including sensory quality, nutritional value and technological quality.The development of technological solutions related to the management, feeding and nutrition of animals, combined with their genetic characteristics, directly influences the composition of the meat and the biochemical changes inherent to the transformation of muscle into meat, which impact the sensory characteristics, nutritional value, losses during preparation or industrialization and the stability of products during shelf life (EMBRAPA, 2022;RAMOS & GOMIDE, 2017).
One of the main challenges of the meat industry (especially the pork industry) is the occurrence of Pale, Soft and Exudative (PSE) meat (TREVISAN & BRUM, 2020); such meat is undesirable because, in addition to being rejected by consumers, it impairs the industrial manufacturing processes, causing considerable economic losses (CALDARA et al., 2012;MOURA et al., 2015;RAMOS & GOMIDE, 2017).The measurement of the initial temperature and pH (at 45 minutes after slaughter -pH 45 ) are an important tool for predicting defects that can compromise the quality of meat because they can be linked to changes in color, texture (tenderness and juiciness, for example) and water retention capacity; these factors related to PSE meat, in addition to industrial yield, shelf life and nutritional value, emphasize the importance of these parameters being monitored and controlled in the meat industry (LIMONI et al., 2017;RAMOS & GOMIDE, 2017;SILVA et al., 2014).
Although, parameters previously described in the literature (initial temperature and pH 45 ) and their importance have been emphasized, there is still a lack of awareness of the potential of using these parameters as a tool for establishing management plans and directions, especially since these factors happen together and they can contribute more or less to specific problems depending on the region.
Thus, this study evaluated the temperature and pH of pig carcasses at 45 minutes after slaughter (pH 45 ) and to estimate the occurrence of PSE carcasses as a function of factors related to the pre-slaughter handling of pigs in a commercial slaughterhouse.

MATERIALS AND METHODS
The study was conducted in a commercial swine slaughterhouse with a daily slaughter capacity of 700 animals under supervision of the Federal Inspection Service (SIF), located in the municipality of Lavras, Minas Gerais.Data collection occurred from December 2021 to February 2022.

Site characterization, adopted practices, and data collection
The data collected and analyzed came from the routine activities developed in the processing plant during the summer, and the initial pH and temperature evaluations were performed on the pig carcasses post mortem.In the slaughterhouse, the animals were kept in covered resting pens with a density of 0.6 m 2 /animal and variable resting duration with free access to water, ensuring that at least 15% of the animals in each pen could drink water simultaneously, as recommended by Decrees 711 and 365 (BRASIL, 1995;BRASIL, 2021).
The animals came from commercial farms and were submitted to a fasting period with free access to water; this information was stated in the health bulletins that accompanied the batches.The animals were loaded into compartmentalized trucks (cage-type trucks) and transported to the slaughterhouse, traveling from 3.5 to 419 km.
The resting and fasting duration were defined according to the time of feed withdrawal stated in the health booklet and the time the animals arrived at the slaughterhouse.In the resting pens, lot separation was maintained according to the origin of the animals.
At the time of slaughter, the animals were led in batches to the spray shower with hyper-chlorinated water (minimum 5 ppm), where they remained for at least 3 minutes and were then individually led to the stunning box that used a syringe.In the stunning box, the pigs were stunned using the electrical stunning method (electronarcosis) and then bled on the bleeding table, with no more than 15 seconds between stunning and bleeding, according to Decrees No. 365 (BRASIL, 2021).
The information of each lot was obtained through a structured form that was previously sent to all producers to assess the following: distance from the farm to the slaughterhouse, date and time of feed withdrawal at the farm, truck specifications, date and time of the start and end of loading, time of farm departure and arrival at the slaughterhouse, and date and time of the start and end of unloading.The forms were returned to the slaughterhouse along with the corresponding load of pigs.

Analyzed variables and sample size
The following independent variables were assessed: A -Transport distance between the farm and slaughterhouse, which was divided into four levels: up to 100 km (n= 765), between 100 and 200 km (n=1936), between 201 and 300 km (n=1340) and over 300 km (n=408); B -Animal transport duration from the farm to the slaughterhouse, which was divided into five levels: up to 3 hours (n=109), between 3 and 6 hours (n=808), between 6.1 and 9 hours (n=2120), between 9.1 and 12 hours (n=1372) and over 12 hours (n=40); C -Loading density of pigs in the truck, which was divided into four levels: below 0.35 kg/m 2 (n = 1391), between 0.35 and 0.40 kg/m 2 (n = 1510) between 0.41 and 0.45 kg/m 2 (n = 1065) and above 0.45 kg/m 2 (n = 483); D -Total fasting duration of the animals until slaughter, which was divided into two levels: up to 24 hours (n=2561) and over 24 hours (n=1888); E -Resting duration of the animals until slaughter, which was divided into five levels: up to 3 hours (n=168), between 3 and 6 hours (n=863), between 6.1 and 9 hours (n=1420), between 9.1 and 12 hours (n=1552) and over 12 hours (n=446); F -Average weight of live pigs, which was divided into four levels: below 100 kg (n=454), between 100 and 110 kg (n=1830), between 110.1 and 120 kg (n=1886) and above 120 kg (n=279); G -Average temperature in the resting stalls, which was divided into two levels: up to 24 °C (n= 2159) and above 24 °C (n=2290); H -Maximum temperature in the resting stalls, which was divided into four levels: below 24 °C (n=190), between 24 and 28 °C (n=1774), between 28.1 and 32 °C (n=1854) and above 32 °C (n= 631).
The initial pH, carcass temperature, and occurrence of PSE were the dependent variables and were obtained by measuring a total of 4449 pig carcasses, which had an average live weight of 109.09± 7.19 kg and came from 138 lots of animals.

Analysis of pH and carcass temperature and estimation of PSE occurrence
Measurements of the pH (pH 45 ) and temperature of the carcasses were performed 45 minutes after slaughter.For the measurement of pH and temperature of the carcasses, a portable measuring instrument with a pH measuring tip combined with a temperature probe (Testo 205, Testo AG, Lenzkirch, Germany) was used, inserted at a depth of 3 cm, perpendicular to the muscle.The measurement was performed in the semimembranosus muscle of the right half of the carcass.

Statistical analysis
Statistical analyses were performed using the Statistical Package for the Social Sciences (SPSS) software, version 20.0.The quantitative dependent variables (pH and carcass temperature) were compared between the levels of the independent variables (transport distance, transport duration, loading density, fasting duration, resting duration, average weight, average temperature in the resting area and maximum temperature in the resting area) with Kruskal-Wallis and Mann-Whitney tests, as the nonnormal distribution of these variables was checked with the Kolmogorov-Smirnov test.
An evaluation of the relationship between the qualitative dependent variable (occurrence of PSE meat) and the independent variables (transportation distance, transportation duration, loading density, fasting duration, resting duration, average weight, average temperature in the resting area and maximum temperature in the resting area) was performed via univariate analysis with the chi-square (χ 2 ) test.
Variables that showed a significant association (with P < 0.02) by the χ 2 test were selected for inclusion in the combined model.The risk was calculated by means of the adjusted odds ratio and its 95% confidence interval for each variable that showed a significant association (P < 0.05) in the logistic regression.
Based on the pH 45 values obtained in this study, the carcasses were classified as normal or PSE.The carcasses were classified as normal when they presented a pH 45 higher than or equal to 5.90 and PSE when they presented a pH 45 lower than 5.90 (BARTON-GADE, 1978;ROSSI, 2022).

RESULTS
The initial (i.e., at 45 minutes) pH (pH 45 ) and temperature values for the carcasses in this study are shown in figure 1.The predominant range of pH 45 values for the carcasses was 6.18 to 6.49 (n=2215), representing 49.78% of the carcasses, while temperatures between 38.08 °C and 40.96 °C corresponded to 64.64% of the carcasses evaluated.
Of all the carcasses evaluated, 306 presented a predisposition to developing PSE meat according to pH 45 values (< 5.90); this number corresponded to an incidence of 6.87% in this study.Table 1 shows the results regarding the effects of the variables on the temperature and initial pH in the semimembranosus muscle of the carcasses.Carcass temperature at 45 minutes varied as a function of distance, and transport duration, loading density, fasting duration, resting duration, and average and maximum temperature of the resting area.There was no difference according to the average weight of the animals.pH 45 values varied as a function of all independent variables studied.
Ciência Rural, v.53, n.12, 2023.Souza et al.Animals transported between 201 and 300 km presented higher carcass temperatures and lower pH 45 values.Regarding the transport duration, the lowest pH 45 were obtained from animals transported for up to 6 hours, and the highest temperatures were obtained from animals transported up to 3 hours.Loading densities lower than 0.35 kg/m² led to higher carcass temperatures and loading densities higher than 0.45 kg/m² led to lower pH 45 values.Regarding the average weight of animals, the lowest pH 45 values occurred in carcasses from animals above 120 kg.Higher temperatures and lower pH 45 values were associated with fasting duration longer than 24 hours.Animals with resting duration up to 6 hours showed lower values for pH 45, while higher temperatures were obtained from the carcasses of animals with resting durations over 12 hours.The average temperature of the resting area also influenced the dependent variables: temperatures up to 24 °C promoted higher carcass temperatures, while temperatures above 24 °C caused lower values of pH 45.
Figure 2 shows the occurrence of meats with different pH 45 values within the same temperature range, indicating that, as an isolated factor, temperature does not determine the predisposition for the PSE.Conversely, the results showed that there was an influence of fasting duration and average temperature in the resting area on the incidence of carcasses with PSE (Table 2).The animals fasted for more than 24 hours had a higher risk (1.41 times higher) of PSE meat, as did those with average temperatures in the resting area above 24 °C (1.52 times higher).The other parameters evaluated did not influence the occurrence of PSE (Table 2).
The transport duration results consolidate these transport distance findings because shorter transport durations of up to 3 hours led to higher temperature and lower initial pH values of carcasses.In addition to transport distance and duration, another variable associated with transport is the loading density.Pigs transported over short distances and at high loading densities during hot weather conditions need more time in the resting area to recover from the stress associated with transport; when undergoing longer transport durations (>3 h) and provided with sufficient space, the animals can acclimate to the transport conditions and partially recover from the stress induced during loading (COBANÓVIC et al. 2021).The highest temperature values in the study were reported in the carcasses of animals transported at loading densities below 0.35 kg/m², lower initial pH values were observed from animals transported at densities greater than 0.45 kg/m².Transporting pigs at low density also increases injuries due to the excess space, as animals may collide with the sides and divisions of the truck, in addition to being at higher risk of slips and falls (DALLA COSTA et al., 2021).
The lowest pH 45 values and highest temperatures occurred in animals with a live weight above 120 kg, the largest weight considered in this study.ELLIS & BERTOL (2001) studied the effects of slaughter weight on meat quality and inferred that heavier pigs have a greater risk of producing PSE meat.Similarly, BERTOL et al. (2015) reported lower initial pH values in heavier pigs.The explanation for this susceptibility lies in the fact that heavier carcasses take longer to cool due to the surface area × mass ratio.
In addition, there is evidence that muscle glycogen content may be higher in heavier animals, increasing the risk of post mortem glycolysis and a faster pH drops.

Occurrence of PSE
The occurrence of PSE meat results from the elevation of the glycolysis rate immediately before and immediately after slaughter, which causes

Variables
--------------Classification--------------------Total-----P value * OR (95%) --------Normal-------------PSE   et al. 2012;DALLA COSTA et al., 2021).The incidence of PSE meat in carcasses from animals fasted for more than 24 hours increased by 37.29% (P < 0.05).These results showed that there is a positive correlation between the increase in fasting duration and loss of meat quality because (in addition to increasing the number of carcasses with pH 45 values below 5.90) there was a decrease in pH values of the carcasses and an increase in temperature, which are the two factors that lead to the occurrence of PSE meat.Prolonged fasting duration can increase fights and damage meat quality, such as leading to PSE meat, since physiological stress can increase the release of hormones that exacerbate muscle activity, resulting in higher rates of glycolysis in the muscle (ABCS, 2014, ABCS, 2016).
Similar results were reported by ACEVEDO-GIRALDO et al. (2020), who evaluated total fasting durations from 3 to 34 hours, with 2 hours allotted for transport, and found that total fasting duration had significant effects on pH values, leading to a reduction in pH 45 values and negative impacts on the color and water holding capacity of the meat (P < 0.01); they concluded that fasting durations should not exceed 12 hours for animal welfare and better meat quality.DRIESSEN et al. (2020) describe the preslaughter fasting duration as a tool to increase muscle pH and decrease the incidence of PSE pork, pointing out that fasting duration less than 18 hours can increase the incidence of PSE meat and duration above 22 hours induced muscle glycogen depletion, increasing the risk of DFD (Dark, Firm and Dry) meat; these findings differ from the present results.Similarly, PANELLA-RIERA et al. (2012) reported that fasting duration of 12 hours increased pH 45 values .In addition, animals subjected to fasting duration of 24-26 hours had meats with a lower rate of exudation, evaluated from drip losses (DRIP).LUDTKE et al. ( 2010) noted that a fasting duration above 24 hours promoted excessive energy expenditure and loss in carcass yield and increased the final pH values (24 hours post mortem), interfering with meat quality.
An average temperature in the resting area above 24 °C before slaughter caused a reduction in the pH 45 value and a consequent increase in the incidence of carcasses predisposed to PSE meat by 48.21%.This result indicated that on warmer days, the incidence of PSE meat has a tendency to increase.These results corroborated the findings of CULAU et al. (1993) andGAJANA et al. (2013), who stated that high ambient temperatures on the day of slaughter decreased the initial pH values and increased the frequency of PSE carcasses.Similarly, COBANÓVIC et al. (2021) reported that meat from pigs slaughtered in summer had lower initial pH values and water holding capacity, which are parameters associated with PSE meat.Pigs are sensitive to high ambient temperatures, as this condition stimulates heat stress due to an impaired thermoregulation mechanism resulting from keratinized sweat glands and a large amount of adipose tissue (LUDTKE et al., 2010;MUN et al., 2022).Heat stress immediately before slaughter accelerates muscle glycogenolysis, increasing lactic acid concentration and producing a rapid decrease in muscle pH while the carcass is still warm, resulting in PSE meat (GONZALES-RIVAS et al., 2020;MUN et al., 2022).
When acute stress occurs just before slaughter, the muscles use more energy reserves, metabolism becomes exothermic and, due to the difficulty of pigs in thermoregulating (i.e., dissipating the heat generated), muscle and body temperature increase.Lower muscle pH with high carcass temperature increases protein denaturation and may result in PSE meat.In addition, high muscle temperatures allow short-term accumulation of lactic acid, which alters protein functionality and increases the proportion of PSE meat (FLORES PEINADO et al., 2020;VERMEULEN et al., 2015).
Maintenance of body temperature in pigs, when exposed to higher temperatures during rest, is achieved by increasing heat loss to the environment and reducing metabolic heat production, a stressor to the animal (DRIESSEN et al., 2020;RENAUDEAU et al., 2011).

CONCLUSION
The initial temperature and pH 45 values of the carcasses were affected by pre-slaughter factors.Furthermore, the fasting duration of the animals and the average temperature of the resting area increased the incidence of PSE meat.

Figure 1 -
Figure 1 -Histograms of pig carcasses (measured from the semimembranosus muscle) at 45 minutes: (A) initial pH values (pH 45 ) and (B) initial temperature values.
-Average temperature in the resting area ( o C) - -Maximum temperature in the resting area ( o C) - Souza et al.&SILVA, 2006).Low pH associated with high carcass temperature leads to greater denaturation of myofibrillar proteins with a consequent reduction in the water retention capacity of the meat(CALDARA et al. 2012).Regarding the distance of transport, the animals transported from 201 to 300 km presented muscles with higher temperature and lower initial pH than the other transport distances.MARZOQUE et al.(2020) reported pH values <5.8 in 11% of carcasses of animals transported over distances between 25 and 65 km and noted that animals transported over distances of 320 km did not present pH values lower than 5.8.In the evaluation of the impact of distances of 45, 430 and 700 km on the initial values of pH and temperature byOCHOVE et al. (

Figure 2 -
Figure 2 -Relationship between initial pH and initial temperature (measured from semimembranosus muscle at 45 minutes after slaughter) in pig carcasses.

Table 1 -
Effects of independent variables on carcass temperature and initial pH (measured from the semimembranosus muscle) at 45 minutes post mortem.

Table 2 -
Occurrence of PSE in pig carcasses as a function of the independent variables.