Pork quality is influenced by many factors along the entire pig production chain. The skeletal muscle content of histidine-containing dipeptides (HCD), consisting of anserine (Ans) and carnosine (Car), has been brought into relation with improved pork quality due to its buffering and anti-oxidative properties. In broilers, dietary histidine (His) supplementation has led to an increase in HCD content with ambiguous effects on meat quality. However, there is only limited information available about the effects of His supplementation on the HCD content and meat quality in pigs. The aim of this research was to investigate the effect of supplementing 0.1% His HCl to growing-finisher pigs for 12 weeks prior to slaughter on the HCD content in the longissimus dorsi (LD) muscle and the effect on carcass and pork quality characteristics. A total of 228 pigs were raised at a commercial farm and were fed the same diet until 12 weeks before slaughter, after which half of the pigs received the His supplementation. On the day of slaughter, 76 control and 75 His supplemented barrow carcasses were randomly selected for carcass and meat quality measurements. A subset of 12 samples per treatment were analyzed for muscle HCD content (Ans and Car). The His supplementation increased the total HCD content in the LD muscle by 16.2% (P = 0.03), attributed to an equivalent increase in the Car content, which was by far the predominant HCD. Ultimate pH was higher in the subset and the total group (P = 0.006 and P = 0.01, respectively) when supplemented with His, with no significant impact on pork water holding capacity in both groups. The color did not differ much between the dietary treatments, with only slightly lower a*-values (less red) for the His supplemented group (P = 0.048 and P = 0.073 in the subset and total group, respectively). The His supplemented group showed a 1.4 mm decrease in fat thickness (P = 0.022) and a 0.9%-point increase in lean meat percentage (P = 0.023) in the total group. Overall, these results indicate that 0.1% His HCl supplementation in feed may increase carcass leanness, meat HCD content and ultimate pH.
There is a broad definition of pork quality, which will depend on specific markets and end-users served.
Pork quality is influenced by the interaction between genetics, farm management, transport, pig processing and carcass chilling (Kurt and Klont, 2010), and finally, storage and handling of fresh meat and derived case-ready and further processed products (Cassens et al., 1975; Andersen, 2000; Kurt et al., 2020). Meat quality is built along the whole supply chain, with synergies and antagonisms between the different attributes (Lebret and Candek-Potokar, 2022). Preslaughter stress and handling of the animals shortly before slaughter is an important factor in the formation of pork quality. High stress levels before slaughter, which lead to a rapid postmortem pH fall, will provoke so-called pale, soft, and exudative (PSE) meat. The reduced water-holding capacity of PSE pork is less attractive to consumers and will lead to economic losses for pig and pork processing companies (Faucitano, 2018; Lebret and Candek-Potokar, 2022).
There is a continuous search in the industry to find solutions to improve meat quality in an effective and economical way. Several nutritional strategies are suggested and used to improve meat quality, such as, e.g., vitamin E and natural antioxidant supplementation to improve meat color stability (Buckley et al., 1995). A possible alternative way of enhancing meat quality might be to increase the histidine-containing dipeptides (HCD) content in pig muscles by supplementing feed additives. HCD is composed of the beta-amino acid [beta]-alanine and the essential amino acid L-histidine (His) and consists in pigs mainly of carnosine (Car) and in minor content of its methylated analogs anserine (Ans) and ophidine (Bolderyrev et al., 2013). HCDs naturally occur in all muscles of vertebrate animals at varying levels where they have anti-oxidative and buffering functions (Boldyrev et al., 2013; Baldi et al., 2021). The HCD muscle content in swine varies and is dependent on, amongst others, gender, breed, and muscle type (Barbaresi et al., 2019; D'Astous-Pagé et al., 2017; Boldyrev et al., 2013; Aristoy and Toldrá, 1998). In addition, the HCD content in meat has been positively related to meat quality parameters such as pH, color, and water retention (Ma et al., 2010; Cong et al., 2017 and D'Astous-Pagé et al., 2017). The relation between His supplementation and the concentration of Car in muscle and effects on meat quality has been described in broilers (Kralik et al, 2015; Lackner et al., 2021). However, there is a a lack of data about these effects in swine. A recent study has shown positive effects of supplementing 0.3% His HCl for 12 weeks to growing-finishing pigs on Car content and meat quality parameters (pH and drip loss) in the longissimus dorsi (LD) muscle (Paniagua et al., 2023).
His is an essential amino acid, which means that pigs are not capable of synthesizing this amino acid themselves and thus, it must be supplied through the feed. In the last few years the crude protein content of swine diets has decreased gradually, mainly for environmental and animal health reasons (Wang et al., 2018); hence, the supply of essential amino acids such as His is decreasing as well. The increased availability of unbound amino acids such as lysine (lys), threonine, methionine, valine, tryptophane, and more recently isoleucine, leucine and His play a key role in keeping performance high despite the lowering protein levels in the diets. In practice, levels of His seldomly go below the presumed requirement for fattening set at 0.32 standardized ileal digestible (SID) His/lys (CVB, 2023). However, a continuous reduced supply of His could also impact the muscle HCD content and influence pork quality consequently. A recent dose-response trial during 14 days (7 to 11 kg body weight) using weaned piglets feeding His slightly below and above NRC recommendations (0.26, 0.32, 0.38, 0.43, 0.49 SID His/lys) did not find an effect on performance but found that a level between 0.35 and 0.41 SID His/lys maximized concentrations of the measured HCDs such as Car in LD, hemoglobin in plasma and trypsinogen in pancreas (Cheng et al., 2023). These results suggest that the animal can limit performance loss by compensating His deficiency from HCDs such as Car. Indeed, earlier research by Barbaresi et al. (2019) compared fast-growing Ross 308 birds with slow-growing Sasso T451 birds and observed a higher HCD level in breast and thigh muscle in slow-growing animals compared to fast-growing animals, suggesting that the available His is privileged for protein accretion (growth) above HCD storage. In other words, the continuous decline in supply of His via low CP diets, may not impact performance, but could reduce levels of HCD available in the skeletal muscle and reduce meat quality traits.
The aim of this research was to investigate the effect of supplementing 0.1% His HCl to growing-finisher pigs for 12 weeks prior to slaughter on the content of HCD in the longissimus dorsi (LD) muscle and the potential improvement of carcass and pork quality characteristics.
Materials and methods
Experimental design, animals, and feeds
The trial was executed at a commercial pig farm in The Netherlands (5476 VK, Vorstenbosch, The Netherlands). In total 288 pigs, 90% barrows and 10% gilts (Topigs Norsvin 70, Norwegian Landrace x purebred York-Z), specifically selected for a premium meat quality program (Robusto; Vion Food, Boxtel, The Netherlands), which is characterized by higher intramuscular fat, specific fat coverage, consistency, and lower drip loss levels than the average slaughter pig in the Netherlands, were randomly allocated to two treatments. The pigs were fed the same commercial diets, containing calculated SID His/lys ratios between 0.35–0.36 (Voergroep Zuid, Deurne, The Netherlands), until 12 weeks prior to slaughter. The two treatments consisted of a control (CON) group and a His supplemented group (HIS+), in which 0.1% His HCl (Bestamino, CJ Bio, L-His content min. 72%, ID Nr. 3c352 (EU regulation 2020/1090)) at the expense of wheat was added to the feed for the remaining period until slaughter. The basal diet was based on wheat, barley, corn and soybean meal and contained no added unbound L-His. The formulated SID His/lys ratio was 0.36 and 0.45 for the CON and HIS+ group, respectively. The formulated nutrient composition of the CON and HIS+ diets can be found in Table 1.
At the day of slaughter, all pigs from both groups were transported to the pig processing plant of Vion Food in Boxtel, The Netherlands. After slaughter, hot carcass weight was determined and 76 chilled carcasses of the CON group and 75 chilled from the HIS+ group were randomly selected (only barrows) to determine meat quality parameters on LD muscle taken from the shoulder side of the left carcass middle.
Assessment of meat quality
On all chilled 151 carcasses from the CON and HIS+ group, pork and carcass quality were determined by measuring ultimate pH, drip loss, color, marbling score, fat and muscle thickness and lean meat percentage. The ultimate pH was measured 18 hours postmortem (MPI pH meter: Meat Probes Inc. Topeka, Kansas, USA). Drip loss (%) was determined by measuring the weight difference of the LD sample after 48 hours storage in a meat tray. Color (L*-, a*-, and b*-values) were determined using a Minolta colorimeter (Konica Minolta CR-400 with 8 mm measuring aperture). Marbling score was visually assessed by an experienced evaluator based on the NPPC pork marbling standards, which depicts seven grades from 1.0 (devoid) to 6.0 and 10.0 (abundant). Fat and muscle thickness in mm and lean meat (%) of the carcasses were determined using a fiber optic meat probe (CGM, Capteur Gras/Maigre, Syleps, Lorient, France). Additionally, twelve randomly selected LD samples per group (15%) were consequently frozen, transported and analyzed for Car and Ans (mg/100 g meat). HCD content (Car and Ans) was determined using HPLC as described in Barbaresi et al. (2019).
Statistical analysis
Data was analyzed with IBM SPSS statistics (Version 29.0.1.0). Animal was the experimental unit. Statistics were performed on two datasets. The first dataset contained all 151 samples (CON, n=76 and HIS+, n=75) which were analyzed for the different carcass and meat quality parameters as described above. The second dataset consisted of the subset of 24 samples (CON, n=12 and HIS+, n=12) which were additionally analyzed for HCD content. Both datasets were first tested for normality and homogeneity of variances to decide about the use of a parametric or non-parametric statistical analysis. If both tests were not significant then the data were subjected to an ANOVA test. If either normality or homogeneity of variances were violated, then the Mann-Whitney U test was used to test the effect of His supplementation. Statistical significance was assessed at P < 0.05, a P-value between 0.05 and 0.1 indicates a statistical trend.
Results and discussion
Feed
The results from the measured feed composition are shown in Table 2. The values are in accordance with the calculated values in Table 1. In the HIS+ group 0.08% extractable His was found, corresponding to about 0.1% His HCl, proving dosing of the His HCl was done properly.
HCD and meat quality traits
The results in Table 3 show that the increased dietary His levels in the HIS+ group resulted in a significant increase in Car and total HCD content in the LD samples (P < 0.05) and a trend for an increased Ans content (P < 0.1). This corresponded to a relative increase of 16.4% and 16.2% for Car and total HCD respectively (Fig. 1). Car is the predominant HCD, representing > 95% of the total HCD content. A higher Car content in pork meat over Ans has been described earlier by Aristoy and Toldrá (1998).
The HCD content values found in the LD muscle from CON and HIS+ animals in the present study are actually higher than the values obtained by Paniagua et al (2023), although the analytics were done in the same way, on the same muscle part, by the same laboratory and that CON pigs in our trial had a comparable SID His/lys ratio in their feed, i.e. 0.36 vs 0.37. Paniagua et al. (2023) found in their control group a LD muscle Car content of 269 mg/100 g versus 307 mg/100 g in the treatment group. On top, the treatment group of the current study was only supplemented with 0.1% His HCL vs 0.3% His HCl in the Paniagua study for the same period (12 weeks). This large difference in HCD content could be explained by natural variation within swine, gender and genetics. Furthermore, the pigs in the present study were selected for premium meat quality properties and could therefore already contain higher base values of HCD. Nevertheless, there was still a significant increase of 16.4% in Car content in the HIS+ group, which is slightly higher compared to Paniagua et al. (2023) who observed an increase of 14%. Paniagua et al. (2023) did also not observe an effect of His supplementation on the Ans content in LD muscle. Their Ans values in the CON samples (5.36 mg/100 g) were threefold lower the Ans values in the present study.
The corresponding carcass and meat quality characteristics from the subset are shown in Table 4. There was a significant (P = 0.006) increase in ultimate pH and a decrease in the a* color score due to His+ supplementation. The hot carcass weight, fat and muscle thickness as well as the other parameters did not differ among the groups.
The higher ultimate pH of the HIS+ group compared to the CON group is positive for meat quality (at least in this range of pH values) as a higher ultimate pH usually coincides with lower drip loss and a darker, more pinkish meat color. However, the significant increase in ultimate pH in this study only resulted in a numerical reduction in drip loss from 1.63% to 1.41%. Paniagua et al. (2023) observed a significant reduction in drip loss from 2.49% to 1.96% for their CON and HIS+ group in combination with a significant increase in ultimate pH from 5.58 to 5.68 for the CON and HIS+ samples, respectively. The drip loss values in our study were lower than the values in the study of Paniagua et al. (2023). Drip loss measurements were not done in the same laboratory, and thus caution is needed when comparing this parameter between studies. The meat samples in our study were from a premium pork quality program, which means that the water-holding capacity of the CON group samples might already be at a higher level, and upgrading pork with an already low drip loss is likely more challenging.
We observed no significant changes in color values for L* and b* in the subset, for both parameters the values were numerically lower for the HIS+ group. There was a significant (P = 0.048) difference for color a* values, with a higher value for the CON group compared to the HIS group. In general, a lower L* and a higher a* are preferred for pork products. The L* value is a measurement for pork lightness with higher values denoting paler meat color. A higher ultimate pH usually coincides with lower L* values and thus darker pork. The lower a* value of the HIS+ group should coincide with slightly less pinkish/red pork, however the magnitude of the difference suggests that this will be hardly noticeable for the human eye. Lackner et al. (2021) did not find significant differences in color values in broiler breast filets from birds fed different His levels at different slaughter ages. Also, no differences were observed in visual marbling levels between the test groups in the subset.
The carcass and meat quality data from the full dataset (n=151) of CON and HIS+ carcasses and LD samples are shown in Table 5. In contrast to the subset, there were significant differences between the groups with regard to fat thickness and lean meat percentage. A 1.4 mm lower (P = 0.022) fat thickness was observed for the HIS+ compared to CON group, resulting in an almost 1%-point higher (P = 0.023) lean meat percentage. It should be noted that similar differences (albeit not significant) were found in the subset.
The ultimate pH of the HIS+ group was also significantly higher (P = 0.01) than the CON group, which is in line with the results of the subset. According to Baldi et al. (2021) there is a relationship between the muscle content of Ans and Car and its postmortem metabolism. An increased level of HCD will increase the buffering capacity of skeletal muscles, leading to a slower postmortem pH decline, thus explaining the slower and reduced extent of muscular acidification of muscle after slaughter and significant differences in ultimate pH values, corresponding to the findings in our research. No significant effects of His supplementation on drip loss and marbling were observed in the full dataset. Overall, these results are in contrast with literature showing that higher HCD content improves meat quality parameters such as drip loss and color in pork (Ma et al., 2010; D'Astous-Pagé et al., 2017). Cong et al. (2017) studied the effect of Car supplementation in broilers and found a significant improvement in meat quality, which could be partially ascribed to increasing antioxidant capacity and decreasing lipid peroxidation status, leading to slower rate of postmortem pH fall and significantly less drip loss. We did find that ultimate pH was increased significantly in the subset and full dataset. However, drip loss was only numerically lower in the subset and showed similar values in the full dataset. The lack of response for drip loss could be related to the fact that the pigs used in this trial already had very low drip loss values and that further improvements are more difficult to obtain.
The color values in the larger dataset showed the same tendencies as the smaller dataset. Both L* and b* values were non-significantly different (P = 0.332 and P = 0.178), but numerically lower in the HIS+ compared to the CON, whilst the a* value showed a trend (P = 0.073) for a higher value for the CON versus the HIS+ group. Overall, the effect on color by His supplementation seems minimal. Cong et al. (2017) studied the effect of Car supplementation on the meat quality in broiler breast muscles and did not find any significant differences in ultimate pH, but a significantly slower pH fall in the muscle after slaughter. Cong et al. (2017) also used a Minolta color meter and did not find significant differences in L* and b*-values, but the a*-value significantly increased in broiler breast with increasing levels of dietary carnosine supplementation.
Conclusions
Supplementing 0.1% His HCl during the last 12 weeks before slaughter significantly increased the muscle Carnosine and total HCD content by 16.4% and 16.2%, respectively. The ultimate pH was also increased significantly with, however, no significant impact on pork water holding capacity. The lack of response for drip loss could be related to the fact that the pigs used in this trial already had very low drip loss values. The color did not differ much between the dietary treatments, with only a slightly but significantly lower a*-value (less red) for the His supplemented group. The His supplemented group showed a significantly lower fat thickness and higher lean meat percentage, which indicates that His supplementation could yield leaner carcasses.
Practical importance
Histidine supplementation showed significant increases in HCD content and in ultimate pH of premium quality pork. The effects on other pork quality parameters were limited. Histidine supplementation could be one of the tools to optimize carcass and meat quality across the entire pig production chain and warrants further investigation in regards to dose and timing of supplementation.
Zusammenfassung
Auswirkungen von L-Histidin-Supplementierung im Futter auf die Schweinefleischqualität bei Mastschweinen
Esra Kurt, Bart Matton, Ronald van Kol, Bas Phillipse, Stefaan De Smet und Ronald E. Klont
Qualität von Schweinefleisch | Histidin | Histidin-haltige Dipeptide | Carnosin | Schweinemast
Die Qualität von Schweinefleisch wird entlang der gesamten Produktionskette von vielen Faktoren beeinflusst. Der Gehalt an histidinhaltigen Dipeptiden (HCD) im Skelettmuskel, bestehend aus Anserin (Ans) und Carnosin (Car), wird aufgrund seiner puffernden und antioxidativen Eigenschaften mit einer verbesserten Fleischqualität bei Schweinen in Verbindung gebracht. Bei Masthähnchen führte die Ergänzung von Histidin (His) im Futter zu einem Anstieg des HCD-Gehalts, mit uneindeutigen Auswirkungen auf die Fleischqualität. Bezüglich der Auswirkungen einer Histidin-Supplementierung auf den HCD-Gehalt und die Fleischqualität bei Schweinen liegen nur begrenzte Informationen vor. Ziel dieser Studie war es, die Wirkung einer 12-wöchigen Supplementierung mit 0,1% Histidin-Chlorid (HCl) auf den HCD-Gehalt im Musculus longissimus dorsi (LD) von Schlachtschweinen sowie auf Schlachtkörper- und Fleischqualitätsmerkmale zu untersuchen. Insgesamt 228 Schweine wurden in einem kommerziellen Betrieb aufgezogen und erhielten bis 12 Wochen vor der Schlachtung das gleiche Futter. Danach wurde die Hälfte der Tiere mit dem Histidin-Supplement versorgt. Am Schlachttag wurden 76 Kontroll- und 75 mit Histidin supplementierte Schweinekadaver stichprobenartig für Schlachtkörper- und Fleischqualitätsmessungen ausgewählt. An einer Teilgruppe von 12 Proben pro Behandlung wurde der HCD-Gehalt (Ans und Car) im Muskel analysiert. Die Histidin-Supplementierung erhöhte den gesamten HCD-Gehalt im LD-Muskel um 16,2% (p = 0,03), was auf einen ebenso starken Anstieg des Carnosin-Gehalts zurückzuführen ist, das mit Abstand das dominierende HCD darstellt. Der End-pH-Wert war in der Teilgruppe und der gesamten Gruppe bei Histidin-Supplementierung höher (p = 0,006 bzw. p = 0,01), ohne signifikanten Einfluss auf die Wasserhaltekapazität des Fleisches in beiden Gruppen. Die Farbe unterschied sich nicht wesentlich, lediglich die a*-Werte (weniger Rot) fielen bei der mit Histidin supplementierten Gruppe etwas niedriger aus (p = 0,048 und p = 0,073 in der Teilgruppe bzw. der gesamten Gruppe). Die mit Histidin supplementierte Gruppe wies eine 1,4 mm geringere Fettdicke (p = 0,022) und einen 0,9%-Punkte höheren Anteil an magerem Fleisch (p = 0,023) in der gesamten Gruppe auf. Insgesamt deuten diese Ergebnisse darauf hin, dass eine Ergänzung des Futters mit 0,1% Histidin-Chlorid den Schlachtkörperfettgehalt senken und den HCD-Gehalt sowie den pH-Wert des Fleisches erhöhen kann.
Authors' addresses
Esra Kurt, and Ronald E. Klont (Corresponding author email: ronald.klont@vionfood.com), Vion, Boseind 15, 5281 RM Boxtel, The Netherlands; Bart Matton, CJ Europe GmbH, Research Center, 60549 Frankfurt am Main, Germany; Ronald van Kol, and Bas Phillipse, Voergroep Zuid, De Poort 3a, 5751 CN Deurne, The Netherlands; Stefaan De Smet, Laboratory for Animal Nutrition and Animal Product Quality, Ghent University, Belgium
Received: 8 February 2024 | reviewed: 4 March 2024 | revised: 8 March 2024 | accepted: 8 March 2024
Schlüsselwörter:
Pork quality
Histidine
Histidine-containing dipeptides
Carnosine
Finishing pigs
Graph: Effects of dietary L-Histidine addition on pork quality in finishing pigs - Gesamter Artikel als PDF inkl. Grafiken
PHOTO (COLOR): Feed formulation - Tab. 1: Formulated nutrient composition of the Control (CON) and Histidine (HIS+) enhanced feed Tab: 1: Nährstoffprofil des Kontrollfutters (CON) und des mit Histidin angereicherten Futters (HIS+) Feed composition - Tab. 2: Measured nutrient composition and lysine and histidine content of the Control (CON) and Histidine (HIS+) enhanced feed Tab. 2: Zusammensetzung sowie Lysin- und Histidingehalt des Kontroll-(CON)- und mit Histidin-(HIS+)-angereicherten Futters Muscle HCD content - Tab. 3: Means and standard deviation of HCD content of randomly selected LD muscle samples taken from pigs fed diets supplemented either with (HIS+) or without (CON) additional histidine. Tab. 3: Mittelwerte und Standardabweichungen des HCD-Gehalts in zufällig ausgewählten Proben des Musculus longissimus dorsi (LD) von Schweinen, die Futtermittel mit (HIS+) oder ohne (CON) zusätzlichem Histidin erhielten. Fig. 1: Percentual increase of carnosine and total histidine containing dipeptides (HCD) levels in LD muscle of pigs fed Control (CON) and Histidine (HIS+) enhanced feed. Source: KURT et al. Abb. 1: Prozentuale Zunahme von Carnosin und dem Gesamtgehalt an Histidin-haltigen Dipeptiden (HCD) im LD Muskel von Schweinen, die mit normalem Futter (CON) und mit Histidin angereichertem Futter (HIS+) gefüttert wurden. Quelle: KURT et al. Carcass and meat quality characteristics - Tab. 4: Means ± standard deviations of carcass and LD meat quality characteristics from randomly selected pigs fed diets supplemented either with (HIS+) or without (CON) additional histidine (same subset as in Table 3). Tab. 4: Mittelwerte ± Standardabweichungen der Qualitätsparameter für Schlachtkörper und Muskelfleisch von Schweinen, die Futtermittel mit (HIS+) oder ohne (CON) zusätzlichem Histidin erhielten. Carcass and meat quality characteristics - Tab. 5: Means ± standard deviations of carcass and LD meat quality characteristics from pigs fed diets supplemented either with (HIS+) or without (CON) additional histidine (full dataset). Tab. 5: Mittelwerte ± Standardabweichungen der Schlachtkörper- und LD-Fleischqualitätsmerkmale von Schweinen, die Futtermittel mit (HIS+) oder ohne (CON) zusätzlichem Histidin erhielten.
By Kurt Esra; Matton Bart; Kol Ronald van; Phillipse Bas; Smet Stefaan De and Klont Ronald E.
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