Blood serum metabolites
The existing literature related on blood serum metabolites in pigs is scarce and reference laboratory values may be useful to detect diseases in pigs based on values derivate from healthy pigs complying different factors such as age, breed, sex, diet, geographical habitat, and methods of sample collection and laboratory measurement (42). Nevertheless, baseline blood metabolite values and how these are affected when pigs are fed different levels of protein and energy at a farm level at a specific age to evaluate the nutritional value of a diet are not available.
Differences in levels of protein and energy in diets had an effect on the blood metabolite profile in finishing pigs at 20 weeks of age. Serum total protein (45–70 g/L) is the sum concentration of all individual serum proteins (42). Prior studies suggested that serum total protein could be used as an indicator of the adequacy of dietary protein content in pigs (43). Several reports have shown that grower pigs fed insufficient crude protein diets up to 60% of the lysine requirements cause a decrease of serum total protein concentration in blood, which may persist during the subsequent finisher phases (13, 44). Moreover, Zeng et al. (11) observed a decrease of serum total protein concentration in blood when grower pigs were fed 65% of the lysine requirements without modifying the levels of crude protein between diets. However, no differences were observed between HCP and LCP diets in finishing pigs in the present study. This result agrees with that of Regmi et al. (12) who did not observe differences in serum total protein concentration in finishing pigs fed insufficient (0.32%), adequate (0.60%) or excess (0.87%) SID lysine diets, and they suggested the pig age difference as the cause to explain the inconsistency in this aspect. Therefore, finishing pigs may be able to show an homeostatic control of serum total protein concentration in blood besides the dietary protein content (12). Moreover, finishing pigs have already reached the maximum protein deposition (45–47) and their metabolism may not be focused on protein turn-over contrary to early stages of the grower-finisher period. Then, serum total protein may be a good indicator of protein synthesis during the growing phase, but not in the finishing phase when pigs are fed insufficient levels of crude protein up to approximately 85% of the requirements such as the present study.
Serum albumin (19–40 g/L) is the most abundant circulating protein found in serum, which accounts for the 60% of the total plasma proteins (42). Albumin plays a major role as a modulator of the plasma colloid osmotic pressure, participating in the transport of hormones, enzymes, fatty acids, metal ions and medicinal products (48). Albumin is considered being a sensitive indicator of protein synthesis capacity of the liver (49) and dietary protein nutrition in pigs (43). However, albumin concentration did not differ in finishing pigs between dietary treatments in the present study, while previous research observed differences between low and high crude protein and amino acids diets when fed in growing pigs (13, 14, 44). Pigs fed insufficient crude protein diets up to 60–80% of the requirements have a reduced albumin concentration in blood (13, 14, 44) leading to a hypoalbuminemia in cases of severe protein restrictions with levels far below 60% of the pigs’ requirements (50–52); while hyperalbuminemia was reported with high crude protein diets (53). Moreover, Regmi et al. (12) observed reduced plasma albumin concentration when finishing pigs were fed a 0.32% SID lysine diet during 4 weeks, but no differences were observed between pigs fed 0.60 and 0.87% SID lysine diets, which are more similar to the SID lysine levels of the dietary treatments of the present study. Thus, serum albumin concentration could be a good indicator of protein synthesis during the growing phase, but not in the finishing phase where differences between levels of crude protein dietary treatments are not observed if pigs are not subjected to a severe low protein diet for a prolonged time.
Serum urea nitrogen is the principal end product of protein catabolism. Amino acid catabolism results in ammonia that is transformed into urea, which it will be transported via blood circulation to the kidney for filtration and posterior excretion via urine (54). Serum urea nitrogen has been used previously as a predictor of efficiency of dietary crude protein utilization (15, 17, 55) and dietary amino acid requirements (11, 12, 16). Serum urea nitrogen was increased in pigs fed the HCP diet in the present study. This finding is in agreement with previous literature that observed increased SUN concentration due to excessive consumption of protein, which was then inefficiently used (14, 15, 17). The same happens when pigs are fed lysine insufficient diets up to 60% of the pigs’ requirements due to an increase of extra free amino acids which will be catabolized through deamination, after the first limiting amino acid is used up (11, 12, 16). The latter was not observed in pigs fed the low crude protein dietary treatment probably because the crude protein and amino acid levels were not so restrictive for finishing pigs, and because amino acids were formulated based on the ideal protein concept (56). Coma et al. (16) stated that the feeding time required to obtain a constant SUN concentration after changing the diet is 3 days. Overall, SUN may be a good indicator of protein efficiency in pigs. Nevertheless, it is worth mentioning that SUN may be altered due to an increase of energy intake (57) or by the use of low crude protein diets supplemented with synthetic amino acids as they could decrease the cation:anion ratio obtaining lower SUN concentrations (16).
Serum creatinine was lower when pigs were fed the LNE diet versus the HNE diet. This result is contrary to that of Hong et al. (17) who found that finishing pigs fed a low energy diet (13.65 MJ/ME) had higher levels of creatinine than pigs fed a high energy diet (14.07 MJ/ME) because of more lean tissue and less fat depositions after 13 weeks of changing the diet. Creatinine (90–240 µmol/L) is produced as the result of normal muscle metabolism (42). Thus, serum creatinine has a positive correlation with total and striated muscle (58, 59). Nevertheless, the discrepancy between the present study and that of Hong et al. (17) could be attributed to the duration of the feeding regime. The present study fed the diets for 10 days to finishing pigs. The difference in added fat and dietary protein/energy ratio between the low and high energy diet could have had a role in changing the protein/lipid metabolism, increasing the serum creatinine concentration due to an increase of carcass fatness and a reduction of carcass leanness (60, 61). This change in the protein/lipid metabolism might be exacerbated based on the protein deposition curve over the grower-finisher period, which declines in finishing pigs (45–47).
Glucose (4.7–8.3 mmol/L) is the principal source of energy for animal cells (42). Pigs fed the LNE diet tended to have lower serum glucose concentration than pigs fed the HNE diet in the present study. Previous literature did not find differences in serum glucose concentration when diets had the same energy levels (11–13) which is consistent with the results obtained in the present study. Nevertheless, Mule et al. (14) found decreased serum glucose concentration at the end of the finisher phase in pigs fed a HCP diet. The same authors attributed this difference due to the decreased amount of carbohydrates in the HCP diet, as glucose is the most important product of carbohydrate metabolism. However, this finding was not observed in the present study, maybe due to a short term feeding regime of the dietary treatments in comparison to that of Mule et al. (14).
Triglycerides are involved in lipid metabolism and the major source of lipid comes from the diet. Pigs fed the HNE diet showed increased serum triglycerides concentration than pigs fed the LNE diet. Lipids are a concentrated energy source and supplemental fats and oils may be added to swine diets to increase energy density of the diet (18). Fats and oils are highly digestible energy sources for pigs and apparent total tract digestibility of lipids is increased with age (18). Therefore, increased serum triglycerides concentration in pigs fed the HNE diet might be explained because of the increased added oil in the diet compared to no added oil in the low net energy diet, affecting the lipid metabolism. Mule et al. (14) reported lower serum triglycerides concentration in finishing pigs fed a HCP diet which seems to be consistent with other research that found a significant correlation between dietary protein restriction and body fat deposition (62). This outcome was not observed in the present study, maybe because of the short term feeding regime of the dietary treatments and/or that HCP diet had a higher inclusion of oil than low crude protein diet.
Cholesterol (1.4–3.1 mmol/L) it is also involved in lipid metabolism and it is derived from dietary sources and synthesized in vivo from acetyl-CoA in the liver as the main site (42). Pigs fed the LCP diet had numerically high serum cholesterol concentration. Previous literature reported a hypercholesterolemic effect when pigs are subjected to dietary protein restriction (19) and amino acid deficient diets (12, 14). It is not clear yet the exact mechanism for the increased serum cholesterol in LCP diets, although earlier studies indicated the insulin/glucagon ratio as an early metabolic index of the effect of dietary proteins and serum amino acids on serum cholesterol levels (63), or that serum albumin acts as a shuttle to enhance cholesterol efflux from cells (64). However, no differences were observed in serum cholesterol concentration between HCP and LCP dietary treatments in the present study. This discrepancy could be attributed to the high added fat in the HCP diet or because of the amino acid levels were not as restricted as previous studies (12, 14). Nevertheless, pigs fed the LNE diet had lower serum cholesterol concentration than pigs fed the LCP diet, which could be related to the no added fat in the LNE diet.
Taken together, the blood serum metabolite profile might be useful towards understanding dietary imbalances and SUN seems to be the best indicator in terms of protein efficiency. Nevertheless, further work needs to be done to establish standard intervals of each serum metabolite to understand whether pigs are fed over or below nutrient requirements, taking into account the pig genetics (14, 55). Further research could also be conducted to determine the separate effect of crude protein and amino acids in the blood metabolite profile.
Volatile fatty acids profile
No differences in the total VFA profile were observed between the dietary treatments. Previous literature reported that a reduction of crude protein in the diet reduced the production of short- and branched-chain fatty acids in manure in growing and finishing pigs (20, 65, 66). Nevertheless, the VFA profile changes from the colon to the manure after a few days of storage (67), so faecal VFA profile might not be comparable to the manure VFA profile. Moreover, the non-difference in total VFA might be explained by the fact that finishing pigs have already a developed gastrointestinal tract with a high fermentation capacity that makes it difficult to observe differences between the dietary treatments of the present study. On this line, a recent study reported that the increased body weight and age of the pigs resulted in an improved digestibility of dietary fiber fractions, which will influence the VFA production (68). In addition, previous literature also showed that the concentration of VFA in faeces is positively correlated with the apparent total tract digestibility of insoluble dietary fiber and cellulose, which are the best factors for predicting faecal VFA concentration (69). Therefore, the difference observed in the BCFA between pigs fed the LNE and HNE diet could be related to the added fiber in the LNE diet. Soybean hulls have a great fermentation capacity (69), which could have produced a shift in the VFA profile reducing the BCFA production by the microbial population. On the same page, the differences observed from pigs fed the control dietary treatment could be attributable to a low fiber content which was an unexpected outcome. Ziemer et al. (70) reported that changes in the faecal VFA profile are influenced by the percentage of cellulose added to the diet, while the level of crude protein in the diet influences the manure VFA profile. Therefore, it could be useful to add the analysis of manure as an indicator of dietary imbalances. Moreover, the non-analysed microbiome gastrointestinal tract could have given a clue of the VFA profile (20).
Thus far, the evidence presented in the present study suggests that blood metabolite and VFA profiles may be affected even before a visual change in growth performance is observed. Furthermore, the changes observed in the blood metabolite and VFA profile could give us a clue whether pigs are fed over or below the nutrient requirements and the high or low inclusion of some nutrients, such as fat or fiber in the diets. Ultimately, the early detection of blood metabolites and/or VFA changes in pigs could improve performance and health and prevent economic losses for pig producers because of inefficiency use of the diets or changes in carcass composition.