Dietary Supplementation with Aqueous Extract of Talinum triangulare at Different Phases of Broiler Production and Effect on Growth, Serum Biochemistry and Carcass Quality

The high cost of conventional feed ingredients, sanctions against the use of chemical growth-promoters, the problem of economic loss and health issues associated with excessive fat deposition are critical concerns in broiler production. Also, the use of antibiotics and anabolic steroids as growth promoters in the livestock industry is facing serious criticism because of problems associated with microbial resistance to drugs, the issue of drug residues in meat and issues that pertain to health of consumers (Alebachew et al., 2016). Moreover, hypercholesterolemia is a major risk factor for heart disease, xanthomatosis, Azheimer’s disease and atherosclerosis (Olson, 1998).

In the poultry industry, the production of broiler chickens with excessive body fat is currently seen as a significant economic problem (Ma et al., 2008). The deposition of abdominal fat in broilers results in increased feed cost and poor final meat quality (Assaf et al., 2004). Researchers are currently looking for natural ways of further reducing the cholesterol level in poultry especially for the diet of populations at risk or “cholesterol shy” persons (Ekpo et al., 2007). This work was targeted at finding a natural alternative to the use of steroids and antibiotics that will achieve desired meat quality in broiler meat.

Talinum triangulare (waterleaf) is a cosmopolitan herb belonging to the family Portulacea (Swarna and Ravindhran, 2013; Brasileiro et al., 2015). Talinum triangulare is a low-cost, natural feed ingredient and growth promoter (nutraceutical agent) (Nya et al., 2010). It is reported to possess numerous medicinal values (Swarna and Ravindhran, 2013; Brasileiro et al., 2015). In a previous study we observed that inclusion of aqueous extract of Talinum triangulare (TT) at a concentration of 1000mg/liter in drinking water of broilers reduced serum cholesterol and the level of fat deposition in the body. However, the phase of production at which such an inclusion would yield an optimum result is yet unknown. This study sought to find out the effect of TTE on the growth parameters, serum lipid and protein profiles and abdominal fat deposition when supplemented at different phases of broiler production.

 

Location
 
The study was conducted in the Research Farm of the Department of Animal Health and Production (AHP), Faculty of Veterinary Medicine, University of Nigeria, Nsukka, Enugu State of Nigeria.
 
Experimental animals and design
 
A total of 180-day old commercial broiler chicks (Arbor Acres strain) hatched by Agrited Nigeria Ltd., were purchased at day-old from Onyinyechi Agro-Veterinary Services, Nsukka.  The study lasted for 11 weeks after one week of acclimatization and brooding which followed standard practices. On day one, the chicks were randomly allocated to four treatment (T) groups A, B, C and D. Each group was further separated into 3 replicates; each replicate housed 15 chicks. Diets of broilers were supplemented with Talinum triangulare extract (TTE) in drinking water from week 2 to 12; 2 to 7 and; week 7 to 12 weeks of life for treatment groups A, B and C, respectively. Group D served as the negative control and received plain water without TTE. Feeds for all replicates were isocaloric and isonitrogenous and water was provided ad libitum.

Each replicate was housed in a separate deep litter pen located in the same poultry house to ensure an identical prevailing environment. Dry wood shavings were used as litter material throughout the experiment; Litter was maintained at 2-4 inches depth which was overturned weekly to prevent caking and minimize microbial growth.

The following drugs and vaccines were used for chemo- and immuno-prophylaxis at different times following manufacturers prescription and through the appropriate routes of administration: Newcastle disease vaccine (NDV i/o), NDV (LaSota), Gumboro vaccine, fowl pox vaccine and coccidiostats.
 
Talinum triangulare extract (TTE) preparation
 
The experimental plant (Talinum triangulare) was sourced from Ogige Market Nsukka, Enugu State of Nigeria. The leaves and succulent shoots of the plant were washed and allowed to drain. It was cut into small pieces and was grinded into a pasty consistency with a grinding machine. One litre of distilled water was used to rinse out from the grinding machine every 1kg of TT that was grinded. The mixture was left for 24 hours. Thereafter, it was turned intermittently for 2 hours and was filtered with a domestic sieve. The filtrate was collected in a plastic container and refrigerated. Fresh extracts were prepared on weekly basis.
 
Determination of the concentration of extract
 
Three glass wares, labelled A, B and C, were washed, dried and weighed with electronic weighing balance (Mettler Toledo®). Using a 5ml hypodermic syringe, 1ml of the extract was aspirated and deposited on each of the glass wares. The wares were placed in a hot air oven and left to dry to a constant weight at a temperature of 60oC. Thereafter, the glass wares containing dried TT matter were weighed and values recorded. For each of the glass wares the concentration of the extract in mg/ml was determined as follows:
 

 
The concentration for glass A, B and C were determined and the average was taken as the concentration of Talinum triangulare.

The extract was added in drinking water of experimental birds at a concentration of 1000 mg/litre of water. The volume of extract to be added per litre of drinking water was thus calculated:
 

 
Blood sample collection
 
After proper restraint, 3 ml of blood was collected from the right jugular vein of each of the birds using a hypodermic needle and syringe and was dispensed into pre-labelled plain test tubes (containing no anti-coagulant) (Surgifriend Medical, England). The tubes were kept in a slanted position and thereafter, centrifuged for 10 minutes at 3000g. Resultant supernatant (serum) was decanted and aspirated into labelled 2 ml capacity sample bottles for serum biochemistry determinations.
               
Parameters measured
 
Growth indices assessed include
 
Feed intake
 
The broilers in groups A-D were fed with standard commercial broiler feed from Top Feed® (Premier Feed Mill Nigeria Plc.). Feed was provided to the broilers twice daily and the remaining feed after 24 hours was collected from feeders of each group and weighed.

 
 
Water intake
 
Fresh water was provided to the broilers twice daily. Water not consumed after 24 hours was collected and measured:
 

 
Weight gain and feed conversion ratio (FCR)
 
The weights of the chicks on day one was measured using a sensitive weighing balance. Subsequently the weights of each group were measured and recorded on a weekly basis. The feed conversion ratio was calculated weekly using the formula:
 

 
Serum lipid profile evaluation
 
The serum lipid profile indices were evaluated using commercially available test kits manufactured by Quimica Clinica Aplicada (QCA), Spain and following procedures as described by Rifai et al., (2008): total cholesterol (enzymatic colorimetric method) was done using the QCA total cholesterol working reagent and assayed using a CHEM5V3 semi-automated blood analyser (Erba Diagnostics, Mannheim Germany); triacylglycerol (TAG) (glycerol-phosphate oxidase enzymatic method) and high density lipoprotein-cholesterol (dextran sulphate magnesium (II) precipitation method). The supernatant after the precipitation was assayed using a CHEM5V3 semi-automated analyser (Erba Diagnostics, Mannheim Germany). The LDL- cholesterol was calculated using the Friedewald’s formula (Warnick et al., 1990); VLDL-cholesterol was calculated as 1/5 of serum triacylglycerol (Rifai et al., 2008).
 
Assay of serum enzyme activity and serum proteins
 
Serum enzyme activity and total protein levels were also assayed using commercially available test kits manufactured by quimica clinica aplicada (QCA), Spain. Alanine aminotransferase (ALT) and Aspartate aminotransferase (AST) (Reitman-Frankel colorimetric method); Alkaline phosphatase (ALP)-(Phenolphthalein monophosphate method) (Colville, 2002), readings were taken using a CHEM5V3 semi-automated blood analyzer (Erba Diagnostics, Mannheim Germany).

Total protein (TP) levels were assayed based on the Biuret method (Johnson, 2008), using the QCA TP working reagent and read with a CHEM5V3 semi-automated blood analyzer (Erba Diagnostics, Mannheim Germany). While the albumin levels were assayed based on the bromo cresol green method and read using the CHEM5V3 semi-automated analyzer at 630 nm wavelength (Johnson, 2008). Globulin levels were calculated by subtracting the albumins levels from the total protein level (Johnson, 2008).
 
Carcass quality
 
Carcass weight
 
At the end of the experiment which corresponded with 12^th week of life, 3 broilers from each replicate were weighed and humanely euthanized. Carcass weights were recorded and their dressing percentages were calculated using the formula:
 

 
Relative weight of edible and non-edible parts
 
After the slaughtering and dressing of 3 broilers from each replicate, the liver and gall bladder, intestine, heart, spleen, gizzard, proventricular and fat around the abdomen and organs were scooped and weighed using electronic weighing balance. The relative organ ratio was calculated using the formula:
 

(Gao et al., 2010).
 
Data analysis
 
The data generated from the experiments were analyzed with descriptive and one-way ANOVA statistics using the computer application- SPSS (version 17.0). Treatment means were compared using least significance difference (LSD) and results were presented using tables, a bar chart and figures.

Growth indices
 
Group A (which was supplemented throughout the experiment) gave the highest feed intake, water intake and average weekly weight gain (Table 1). The results of this study suggest that aqueous extract of Talinum triangulare is a potent phytobiotic in broiler production since it enhanced growth parameters and did not adversely affect the health of broilers. Talinum triangulare has high levels of essential nutrients like minerals (such as calcium, potassium and magnesium), soluble fibres (such as pectin) and vitamins (such as C, α and b tocopherols and b-carotene) which are required for growth and development (Ezekwe et al., 2002). Also, the absence of significant difference (p<0.05) in the growth indices (Table 1) indicate that waterleaf possibly has low level of anti-nutritive factors that could adversely affect broiler growth. Experimental Group B recorded the least value of water intake (Table 1) probably due to the sudden change in diet from plain water to treated water. However, Group B had the best feed conversion ratio.


 
Serum lipid profile
 
Supplementation with TTE significantly (p<0.05) increased the serum HDL-C (Fig 1) and lowered the LDL-C (Table 2a) in the treated groups. John et al. (2007), explained that some plants contain a group of chemical compounds known as phytosterols which being similar to cholesterol, compete with the absorption of cholesterol through the gut thereby reducing the LDL and increasing the HDL level in the blood. Secondly, action of the enzyme Hydroxymethylglutaryl-coenzyme A (HMG-CoA) reductase is the regulatory, rate-limiting and irreversible step in cholesterol synthesis (Rhode et al., 1995). Talinum triangulare extract may also contain some active principles similar to HMG-CoA which like reductase competitive inhibitors may reduce the rate of synthesis of LDL. This is subject to further investigation. No significant difference was recorded for total cholesterol, VLDL cholesterol and TAG (Table 2).



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Serum enzymes and proteins
 
Treatment had no significant effect on liver enzymes and serum proteins (Table 3). This suggests that the extract does not have any significant effect on the liver-hepatocellular integrity and synthetic ability.

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Carcass quality
 
The relative organ weights, carcass and live weights of the broilers showed no significant variation across the groups (Table 4). This is in line with the findings of Iweala (2011) who reported that the weight of rats fed waterleaf supplemented diet did not change significantly. The carcass quality as shown in Fig 2A - 2D revealed that very lean broiler carcasses (minimal fat deposition) were produced in groups 2A and 2C. This is evidenced by shrunken lower abdominal and perinea regions, prominent keel bones  and depleted fat deposits in carcasses from groups 2A and 2C. Fat deposition was marked in groups 2B (treated at starter phase) and 2D (not treated throughout the study). It is thought that the high level of fat deposition in group 2B was probably due to some sort of compensatory fat deposition mechanism following the withdrawal of treatment with TTE during the finisher phase of production. The tissue fat level correlates with the LDL and HDL levels in the blood (Mamonto, 1992). Low density lipoprotein (bad cholesterol) is known to increase fat deposition in body tissues and is implicated in many coronary heart diseases, whereas HDL (good cholesterol) mops up cholesterol from the system (Tymoczko et al., 2002).

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Dietary supplementation with aqueous extract of Talinum triangulare at a concentration of 1000mg/litre of drinking water in broiler diet gives the best results, in terms of production of lean poultry meat with better cholesterol profile (higher HDL and Lower LDL), when given throughout the period of production (week 2 - 12). Supplementation during the finisher phase produced lean poultry meat comparable with the former and may be more economically viable. Talinum triangulare extract does not have any significant untoward effect on production parameters, as a natural growth promoter or a non-conventional feed ingredient.

More research should be carried out to define how much of TTE may replace conventional feed stuff on dry matter basis without negative effect on production parameters. More precise test should be used for the evaluation of the effect of the test plant on immunity of birds. In addition, a preliminary organoleptic evaluation of poultry meat from treated groups is recommended to shed more light in this area.