Trace mineral chelation for sustainable animal nutrition : enhancing zinc availability with L-glutamic acid N,N-diacetic acid

In current farm animal practice, the uncertainty in the availability of zinc (Zn), as affected by dietary and digestive factors, is compensated by calculating gross requirements from net requirements using a worse-case availability factor in the conversion. Consequently, the higher levels of Zn inclusion lead to a reduction in relative efficiency of uptake, as levels fed are higher than Zn requirements. Ultimately, the result of this is an increase in Zn manure which can result in high Zn levels in soil when this manure is used, increasing the environmental impact of farm animals. A novel chelator, L-glutamic acid N,N-diacetic acid (GLDA), is a chelating agent, capable of binding di- and trivalent metal ions. By binding to these metal ions, it potentially provides stability of the complex in the upper gastrointestinal tract, which minimizes the formation of insoluble complexes, thereby improving nutritional bioavailability. This thesis aims to improve our understanding on trace mineral nutrition and determine the potential of using GLDA to increase the availability of minerals in livestock production.In Chapter 2 the impact of GLDA was compared to the well-established chelating agent ethylenediaminetetraacetic acid (EDTA). Previous work in literature showed effects of EDTA on trace mineral retention, but EDTA suffers from low biodegradability and its high chelation strength can be considered to be too high compared to metal transporters in the body. In experiment 1 broilers were fed Zn sulphate with GLDA or EDTA in molar amounts equivalent to chelate the level of Zn added. In experiment 2 the effect of GLDA on a basal diet containing no additional minerals was established. Serum and tibia Zn clearly responded to the increasing doses of dietary zinc with a significant response to the presence of EDTA and GLDA. These results are also indicative of the equivalent nutritional properties between GLDA and EDTA. In experiment 2, zinc levels in serum and tibia were also increased with the addition of GLDA to a basal diet lacking supplemental trace mineral, where serum zinc levels were 60% higher at the 216 mg/kg inclusion level. The study demonstrated that dietary GLDA enhanced availability of Zn.The aim of Chapter 3 was to quantify the reduction of dietary Zn that could be achieved in broilers to obtain the same Zn status. Broiler were fed Zn in a dose response manner with and without GLDA. The results indicated that when GLDA was included in the diet, based on tibia Zn, the same Zn status was achieved with a 19 mg/kg smaller Zn dose while based on serum Zn this was 27 mg/kg less Zn.Chelators are known to have negative side-effects when fed at high levels, for example due to chelation of minerals within the cell walls, leading to cell wall disruption. In order to determine the effects of high GLDA inclusion a dose response with GLDA up to 10000 mg/kg was performed in Chapter 4. The results of this study indicated that there are no negative side effects of GLDA inclusion up to 3000 mg/kg of GLDA/kg feed. The GLDA residue levels in breast tissue indicated that 0.01% of total GLDA absorption is stored in breast tissue. Higher values are found in kidney and liver for the highest inclusion level, indicating that the fraction of GLDA that is absorbed is actively excreted by the animal. The limited absorption of GLDA indicates that the role of GLDA affecting Zn availability takes place within the gastrointestinal tract of the animal, by sustaining solubility during digestive processes.As the Zn load in manure of pigs is greater than that of broilers, the effect of GLDA in piglets was determined in Chapter 5. On top, pigs are a better model for potential human implications than broilers, which is important considering the high prevalence of Zn deficiency in the developing world. GLDA appeared to protect a significant fraction of soluble luminal Zn from being captured by phytic acid and mediated it towards the Zn transport mechanisms in the gut mucosa, thereby promoting higher Zn retention in GLDA-supplemented animals compared to control animals. This led to lower necessity for mobilization of body Zn stores to compensate for endogenous losses in the presence of GLDA, lowering the chance of subclinical or clinical Zn deficiency.In Chapter 6 we discussed the impact of GLDA on Zn availability and retention in farm animals by combining the results of previous chapters with existing literature. The importance of study design when assessing trace mineral availability is discussed. The impact of GLDA on trace mineral availability is discussed by showing the increased retention of Zn when using adding GLDA to diets of animals. The potential impact on the environment by implementing GLDA is discussed and put into context regarding sustainable farming, emphasizing the key role GLDA can play

Metabolomic Analysis of Wooden Breast Myopathy Shows a Disturbed Lipid Metabolism

Myopathies have risen strongly in recent years, likely linked to selection for appetite. For white striping (WS), causes have been identified; but for wooden breast (WB), the cause remains speculative. We used metabolomics to study the breast muscle of 51 birds that were scored for both at 35 days of age to better understand potential causes. A partial least square discriminant analysis revealed that WS and WB had distinct metabolic profiles, implying different etiologies. Arginine and proline metabolism were affected in both, although differently: WB increased arginine in breast muscle implying that the birds did not use this pathway to increase tissue blood flow. Antioxidant defenses were impeded as shown by low anserine and beta-alanine. In contrast, GSH and selenium concentrations were increased. Serine, linked to anti-inflammatory properties, was increased. Taurine, which can stabilize the cell’s sarcolemma as well as modulate potassium channels and cellular calcium homeostasis, was also increased. Mineral data and depressed phosphatidylethanolamine, cAMP, and creatine-phosphate suggested compromised energy metabolism. WB also had drastically lower diet-derived lipids, suggesting compromised lipid digestion. In conclusion, WB may be caused by impaired lipid digestion triggered by a very high appetite: the ensuing deficiencies may well impair blood flow into muscle resulting in irreparable damage

Efficacy of l-glutamic acid, N,N-diacetic acid to improve the dietary trace mineral bioavailability in broilers

Trace minerals are commonly supplemented in the diets of farmed animals in levels exceeding biological requirements, resulting in extensive fecal excretion and environmental losses. Chelation of trace metal supplements with ethylenediaminetetraacetic acid (EDTA) can mitigate the effects of dietary antagonists by preserving the solubility of trace minerals. Lack of EDTA biodegradability, however, is of environmental concern. l-Glutamic acid, N,N-diacetic acid (GLDA) is a readily biodegradable chelating agent that could be used as a suitable alternative to EDTA. The latter was tested in sequential dose-response experiments in broiler chickens. Study 1 compared the effect of EDTA and GLDA in broilers on supplemental zinc availability at three levels of added zinc (5, 10, and 20 ppm) fed alone or in combination with molar amounts of GLDA or EDTA equivalent to chelate the added zinc, including negative (no supplemental zinc) and positive (80 ppm added zinc) control treatments. Study 2 quantified the effect of GLDA on the availability of native trace mineral feed content in a basal diet containing no supplemental minerals and supplemented with three levels of GLDA (54, 108, and 216 ppm). In study 1, serum and tibia Zn clearly responded to the increasing doses of dietary zinc with a significant response to the presence of EDTA and GLDA (P < 0.05). These results are also indicative of the equivalent nutritional properties between GLDA and EDTA. In study 2, zinc levels in serum and tibia were also increased with the addition of GLDA to a basal diet lacking supplemental trace minerals, where serum zinc levels were 60% higher at the 216 ppm inclusion level. Similar to the reported effects of EDTA, these studies demonstrate that dietary GLDA may have enhanced zinc solubility in the gastrointestinal tract and subsequently enhanced availability for absorption, resulting in improved nutritional zinc status in zinc-deficient diets. As such, GLDA can be an effective nutritional tool to reduce supplemental zinc levels in broiler diets, thereby maintaining health and performance while reducing the environmental footprint of food-producing animals.</p