In this study, the deficiency of TM (Zn, Fe, Mn, Cu, Se, I, and Cr) in the NTM diet strongly depressed overall growth performance of broilers, which was more noticeable during the early growth period. This normally can be linked to higher rates of growth, bone development, and metabolism during this period [7]. Based on the results of the current study, the CTM25, CTM50, and ITM treatment groups showed no significant differences on growth performance, but the CTM100 treatment showed better performances versus the ITM and CTM25 treatments. Although there was no difference between CTM50 and CTM100 groups on production performances, only the CTM100 diet increased EPI during the overall growth period compared with equal levels of TM from ITM treatment, which indicated that the complete substitution of inorganic TM sources by CTM could be as an alternative way to improve productive performance. This improvement in broiler performance appears to confirm the improved bioavailability of TM from organic sources [2, 9, 22]. However, other researchers have reported variable responses with the addition of organic TM on growth performance. Zhu et al. [6] also reported that there was no negative impact on growth performance due to the supplementation of the diet with organic TM at reduced levels (30% and 50% of the regular inclusion level). Similar results were also observed by De Marco et al. [12], who found that using organic TM sources (metal chelates of glycine and hydrate) in broiler diet could reduce the supplementation of Zn, Fe, Mn and Cu to 50% of the strain recommendations. On the other hand, Sirri et al. [23] reported that substitution of Zn, Mn, and Cu from inorganic sources by their organic chelates (chelated metals methionine hydroxy analogue) significantly improved body weight gain and FCR of broilers from 0 to 51 d of age. The previous study in Vanaraja chickens also demonstrated the positive effect of CTM-supplemented diets, even at low dietary inclusion level (50%), on FCR during the entire experimental period [24]. The inconsistencies in the efficacy of organic minerals may be attributable to different factors. The breed, differences in diet composition, housing condition, type and dosage of the supplement, the duration of the experiment, and bird characteristics can affect the growth response of broiler chickens to organic mineral supplements.
In this study, in addition to growth parameters, the evaluations of mineral bioavailability and tibia characteristics were within the main purposes of this study. When minerals are added beyond the requirement of the animals, more is excreted because of the reduction in utilization efficiency for that mineral [9]. Decreasing the use of TM in poultry feeds may be a feasible solution for minimizing mineral emissions from poultry farms. In the present study, all CTM levels could efficiently improve Zn and Mn digestibility as compared to inorganic mineral supplementation. Because feeding the diet supplemented with 100% of organic TM produced the higher Zn and Mn absorption than the use of 100% of inorganic forms in the current study, it can be said that providing the same levels of organic TM could lead to higher mineral retention. Several researchers have stated the low excretion of organic TM through the excreta and consequently their high retention rate in broilers [6, 12], laying hens [25] and growing ostriches [15]. Based on the current results, it appears that lower supplementation of TM to 50% of the commercial level could improve the absorption of TM, without compromising the growth performance of broilers as previously mentioned. Increased mineral digestibility in broilers fed the diets supplemented with chelated TM is attributed to the fact that organic minerals may be better absorbed as they are more likely to be stable throughout the gastro-intestinal tract and less prone to antagonisms and interactions with other components of the digesta [11]. The results of this study also showed that broiler chickens fed the CTM-supplemented diets had higher digestibility of P compared to the ITM group. A possible reason for this observation is that chelated form of TM may reduce the formation of complexes between P and other metal ions, and, therefore, increase P absorption in the ileum. Another possible reason to explain the greater AID of P might be the improvement in phytate P utilization due to organic acid released from CTM supplement [15], although the mechanism of action is not well-understood.
Trace minerals appear to play important roles in the growth, development, and maintenance of normal bone. Bone as a complex heterogeneous tissue is responsible for supporting muscle, and therefore there is a close link between growth and development of bone with overall body growth [26]. Tibia morphological measurements, such as tibia length, weight, ash, and mineral contents have been used as indicators for the evaluation of bone status in poultry [2]. In the NTM group, tibia length and Mn and Cu contents were lower compared with the ITM100, CTM50, and CTM100 groups, which suggests that a lower amount of bone components was available in group NTM. In the present study, tibia morphological traits and bone mineral content in the CTM50 group were in the same range as those obtained by chickens in the ITM100 group, which could be due to greater mineral absorption and bioavailability in the CTM50 group. These results are in agreement with the results of previous studies (9, 13, 24], in which a stimulatory effect of organic mineral sources on morphological parameters of bone were observed. The same results were also found by M’Sadeq et al. [2], who indicated that broiler chickens fed 37.5 and 50% of organic yeast proteinate TM premix had no effect on tibia strength, weight, length, and width compared with those fed 75% inorganic TM premix or 100% salt encrusted TM premix on day 38 of age. Increased tibia P content in broilers fed the CTM-supplemented diets compared to those fed the ITM diet in the present study also confirms the hypothesis that organic TM supplementation could reduce interference from agents that form insoluble complexes with ionic TM [24]. This might be also a reason for greater tibia ash content observed in the CTM100 group compared to the ITM group.
The metabolomic analysis of serum in the present study revealed that experimental groups only differed in uric acid concentration, which was higher in the CTM100 group than the NTM, ITM, and CTM25 groups, while it was not different from the CTM50 group. The increase in blood uric acid concentration could be associated with the free radical scavenging capacity [27]. In a previous study where broiler chickens were fed diets that replaced inorganic TM with organic TM, the authors found a significantly higher serum uric acid level [1]. In part, these findings could contribute to the possible reduction of tissue damage by organically complexed minerals.
Malondialdehyde (MDA) is the principal product of lipid peroxidation, and its accumulation can reflect the degree of lipid peroxidation [28]. In contrast, the enzymatic scavengers, such as GPx, SOD, and CAT enzymes, help break up the damage process by removing reactive oxygen species [29]. Therefore, SOD, GSH-Px, CAT, and MDA are often used as effective indicators to objectively reflect the antioxidant status in animals [30, 31]. As the antioxidant status of chickens was assessed, broiler chickens fed with any of the diets exhibited lower serum MDA levels than those fed the ITM diet (Table 7). Accompanied by this decrease in MDA content, all TM-supplemented groups exhibited an increase in serum GPx and SOD activities, while the CAT activity remained relatively constant in these chickens. Feeding diets with CTM at the 25 and 50% of dose level suggested by the strain recommendations (CTM25 and CTM50) also increased the activities of Gpx and SOD enzymes and decreased MDA to levels comparable to those observed in broilers fed diets containing sufficient amounts of inorganic TM (ITM group). This may indicate that organic minerals supplemented at lower levels have higher bioavailability and can exert the same antioxidant capacity than higher levels of inorganic forms. Similarly, in another study, total replacement of high levels of inorganic TM by lower levels (50% and 62.5% of the commercial recommended levels) of organic TM in broiler breeder diets had similar activities of serum GSH-Px and SOD. Additionally, we found that birds fed the CTM100 diet had the lowest MDA content as well as the highest GSH content and T-SOD activity in serum among all of the experimental groups in the present study. Similarly, replacing inorganic TM (Fe, Mn, Zn, and Cu from sulfates and Se from sodium selenite) by equivalent levels of organic TM (metal proteinates and selenium yeast product) improved hepatic Cu/Zn-SOD and GSH-Px activities in commercial pigs [32]. The antioxidant stress protection was stated to be effective only if there are adequate quantities of cofactors such as Zn, Mn, Cu, and Se available [15, 33]. Organic Zn, Mn, and Cu have been reported to increase the synthesis of SOD [14, 34], while organic Se previously enhanced GSH-Px activity [35, 36], which is similar to our study. The use of organic TM might have increased the bioavailability of these minerals and thus reduced the accumulation of reactive oxygen species, which might have improved the antioxidant defense system in broilers receiving the CTM100 diet, as compared with those fed the ITM diet in this study.