By Woke, JA; Diri, M; Leton De-Great, KC; Johnson, NC (2023). Greener Journal of Biological Sciences, 13(1): 8-11.

Effect of Feeding Graded Levels of Vitamin C on Antioxidants and Oxidants in Broiler Chickens

¹Woke, J. A.; ^1*Diri, M.; ²Leton De-Great, K. C.; and ¹Johnson, N. C.

¹Department of Animal Science, Rivers State University.

²Department of Agricultural Extension and Rural Development, Rivers State University.

INTRODUCTION

In young growing animal species, such as broiler chickens their fast growing process imposes some degree of stress on the animals during production, especially at the commercial setting where poultry diets are fortified with high density nutrients to maximize growth. Based on the findings of Herbert et al. (2005) and those of Stahly et al. (2007), one of the nutritional means of supporting the animals for enhanced growth and performance is to fortify poultry diets with antioxidant vitamins, such as vitamin C (Johnson et al., 2019).

The fore-stated is primarily true due to the fact that there are inherent measures that animals, including poultry have developed through which they are capable of avoiding oxidative stress. In other words, animals possess a complex oxidation defense system that are modulated by special molecules known as antioxidants whereas the byproduct of oxidative stress is MDA. According to Hyman (2011), antioxidant molecules are glutathione (GSH), superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GSH-Px). The byproducts of the reactions involving these antioxidants are mainly malondialdehyde (MDA). Therefore, MDA levels in the blood are indicators of the level of oxidative stress an animal has been exposed to.

Amongst the antioxidants, GSH is the major or master antioxidant as it is also responsible in preserving its cohort antioxidants. This has earned GSH being referred to as ‘mother of all antioxidants’, master detoxifier and master of the immune system (Hyman, 2011). SOD catalyzes the dismutation of superoxide radical into oxygen or hydrogen peroxide thereby protecting cellular components from oxidation by reactive oxygen species (ROS). CAT converts hydrogen peroxide to carbon dioxide and also uses hydrogen peroxide to breakdown harmful toxins in the body (Sharma and Dubey, 2005). GSH-Px scavenges and inactivates ROS thereby protecting the animal from oxidative damage by acting as an efficient quencher of ROS (Vangronsveld and Clijsters, 1994). Johnson et al. (2019) demonstrated that vitamins C and E upregulated GSH, SOD, CAT and GSH-Px and simultaneously reduced MDA in the pig. There is paucity of information on these parameters in the broiler chicken. Therefore, the objectives of this study are: to investigate the effects of feeding graded levels of vitamin C on antioxidants and to also evaluate the effects of grade levels of vitamin C on oxidants levels in broiler chickens.

MATERIALS AND METHODS

Experimental site

This study was carried out at the poultry unit of the Teaching and Research Farm, Rivers State University, Nkpolu-Oroworukwo, Port Harcourt. The farm is situated at latitude 4⁰ 48’N and longitude 6⁰ 48’E at the Rivers State University campus.

Animals

One hundred and twenty (120) Agrited day-old chicks were acquired from a reputable commercial poultry dealer in Port-Harcourt, Rivers State. The animals on arrival at the Rivers State University Teaching and Research Farm were brooded to properly pre-condition them to their new environment. The animals by the fourth week were observed to have properly adapted to their environment and thus were randomly assigned into four dietary treatment groups of 30 birds/treatment group with 3 replications of 10 birds/replicate. The pens were properly cleaned and disinfected before the birds’ arrival. Feeders and drinkers were also properly cleaned to also ensure that the animals’ environment were “pathogen-free”. During the brooding period all protocols, including the necessary medications were provided. Animals were fed similar diets from day one through the end of the 4^th week. Water was provided ad libitum. The experiment lasted for 8 weeks and thus animals received their respective experimental diets for 4 weeks.

Experimental Diets

Hybrid feed^TM grower mash were used in the study. In other words, the diets fed to the animals during the last four weeks of the experimental period were similar in all nutrients except their dietary vitamin C levels as: control or treatment 1 (T₁, contained only basal level), treatment 2 (T_2, contained 200mg of vitamin C), treatment 3 (T_3, contained 300mg of vitamin C) and treatment 4 (T₄, contained 400mg of vitamin C)/kg of diet, respectively. The animals were fed these graded levels of vitamin C-based diets for 4 weeks.

Blood Sample Collection

At the end of the study period, birds were killed for blood collection. 9 birds were randomly collected from each treatment group consisting of 3 birds from each replicate of the four treatment groups, respectively. The blood was collected from each bird into treated tubes with ethylene diamine tetra-acetic acid (EDTA) and immediately snap frozen for later antioxidants and oxidants analyses.

Antioxidants

Antioxidants molecules analyzed for were: CAT, SOD, GSH and GSH-Px, whereas, the oxidants analyzed for was MDA. CAT was analyzed for according to the method of Aebi et al. (1974). SOD was analyzed for according to the method of Misra and Fridorich (1972). GSH and GSH-Px were measured according to the method of Agerganrd and Jensen (1982). MDA was measured according to the method of Varsney and Kale (1990).

Experimental Design and Statistical Analyzes

The study was designed and conducted as a completely randomized design (CRD). Data obtained were subjected to analysis of variance (ANOVA) using general linear model (GLM) procedure of SAS. Treatment means were compared using Tukey’s test. Therefore, the model was: Y_ij = µ + X_i + E_ij, where Y_ij = individual observation of the treatment, µ = population mean, X_{i =} effect of the i^th treatment and E_ij = the error term. An α-level of 0.05 was used for all statistical comparisons to represent significance.

RESULTS

The results of the antioxidants’ molecules are shown in Table 1.

Table 1. Antioxidants Status of Broiler Chickens Fed Graded Levels of Vitamin C-based Diets

As depicted in Table 1, there were no significant (P > 0.05) differences in the antioxidant molecules amongst all treatment groups. The results of the oxidants fed graded levels of vitamin C-based diets are shown in Table 2.

Table 2. Oxidant Levels of Broiler Chickens Fed Graded Levels of Vitamin C-based Diets

Means within each row with different superscripts differ significantly (P < 0.05)

For the oxidant MDA, there were significant (P < 0.05) differences amongst the treatment groups. For instances in the MDA concentrations, animals in the T₁ group had the highest level and linearly significantly reduced as the level of vitamin C dietary increased with the T₄ group had significantly (P < 0.05) the lowest level.

DISCUSSION

Vitamin C is an antioxidant vitamin as it is actively involved in the regulation of molecules that scavenge for ROS and convert them to less harmful products; implying that they significantly contribute to the overall health status of the animal (Michael et al., 2010: NRC, 2012). Some of these molecules are GSH, GSH-Px, CAT and SOD whereas the production levels of MDA are indicators of the degree of the effectiveness of the antioxidant vitamin activities on the antioxidant molecules.

Antioxidants protect cells, tissues and animals’ organs by reacting with free radicals (Traber et al., 2011; NRC, 2012). These antioxidant molecules are produced endogenously in the body of the animal, such as GSH and its cohorts in the GSH defense system. In animal nutrition, these antioxidant molecules productions can also be triggered by inclusion of antioxidant vitamins to aid in stimulating their productions in maintaining normal health status of the animal (Bourne et al., 2000). Furthermore, it is worthy of note that antioxidants work synergistically in the animal body to elicit effective defense functions (Ursini, 2000: Johnson et al., 2019). Therefore, one antioxidant molecule synergizes with another team member for the system to be very efficient. SOD is widely perceived as the first level of antioxidant defense probably due to it being more resident in the cytosol (Surai, 2002). It converts superoxide radical into hydrogen peroxide and oxygen. GSH-Px and CAT convert hydrogen peroxide to water. GSH acts as co-enzyme by a variety of crucial life processes, such as detoxification of xenobiotic, removal of hydro-peroxides and other free radicals (Surai, 2002). However, there were no differences in the antioxidant molecules investigated in the plasma concentrations of the birds in all treatment groups in the current study. This probably might be related to the fact that there was no known attack on the animal, including pathogens since the animal environments were maintained at high level of sanitation (NRC, 2012) throughout the experimental period.

Conversely, while we found no differences in the antioxidant molecules amongst treatment groups, those of oxidants, namely MDA were significantly reduced in the T₂, T₃ and T₄ animals compared with the T₁ group. This is a clear indication that the ingestion of the vitamin C-based diets reduced oxidative stress in the animals of T_2, T₃ and T₄ groups. MDA is a major by-product of oxidative stress (Guo et al., 2006). Therefore, the significantly reduced levels of MDA in the T_2, T₃ and T₄ animals compared with the T₁ animals was a further confirmation indicating that the animals that received the vitamin C-based diets experienced lesser activities of oxidative stress compared with the T₁ animals. This observation is in tandem with those of Hyman (2011), Achuba and Otuya, (2006) and Johnson et al. (2019). This is again supported by the findings of improved performance of the animals on the vitamin C-based diets that was part of this present study.

CONCLUSION

The ingestion of vitamin C-based diets had no effect on antioxidant molecules; however, it reduced oxidative stress as evidenced by significant reduction levels of MDA in broiler chickens that ingested the vitamin C-based diets.

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