INTRODUCTION
Antibiotics are microbial metabolites produced by fungi and algae which have low molecule weight and can inhibit the growth of other microorganisms even in low concentrations (Nir and Ve-Senkoylu, 2000). While antibiotics have prevalently been used as growth promoters in animal nutrition, European Community has prohibited the use of antibiotics in animal nutrition as growth promoters from January 1, 2006 (Anonymous, 2005). As a result of the ban of antibiotic growth promoters due to the demands from medicine and consumers; explorations on alternative products have started. Consequently, studies on natural products such as plant extracts have recently gain a great attention (Wenk, 2000).
It is stated that the plant extracts can continuously be used in rations without any need for their removal and that they do not induce any resistance to antibotics (Gill, 1999). At present, phytogenic products can be classified as sweeteners and appetizers without a requirement for the determination of their minimum residue levels (Kamel, 2002). On the other hand, there is a misbelief that "all plant extracts are beneficial since they are natural and organic". For example, the plant "EPHEDRA" was prohibited at the end of the year 2003 since it damages nervous system and leads psychosis, memory loss and even death (Anonymous, 2004). Animals, poultry in particular, are very sensitive to pathogenic bacteria such as Escherichia coli, Salmonella sp. Clostridium perfringens and Campylobacter sputorum. The pathogenic microbial flora in the small intestine compete with host for nutrients while at the same time inhibiting the binding of the bile acids to the pertinent substances, they decrease the digestion of fats and fat-soluble vitamins. This leads to a decrease in performance and increase in disease rate. Antibiotics, which have been used as growth promoters in poultry feed for a long time, improve the growth performance by stabilizing the microbial flora in the intestine and preventing some specific intestine pathogens (Gunal et al., 2006).
Another increasingly important recent natural product is propolis. Propolis is a glue-like substance that honey bees collect from plant seedlings and buds. It is obtained as a result of the biochemical alteration of the resinous materials and plant secretions by the enyzmes secreted from the glands of the bees. It has a colour ranging from dirty yellow to dark brown, a strong and nice odor, is water-insoluble and semi-solid in room temperature (Hepsen et al., 1996; Sahinler, 2000). Since the chemical composition of propolis is highly complex and its composition varies according to the plant, region, season and colony; its colour, odor and medical characteristics vary accordingly (Kutluca, 2003). The composition of raw propolis is generally composed of 50% resin and vegetal balsam, 30% beeswax, 10% essential and aromatic oils, 5% pollen and 5% other organic substances (Kumova et al., 2002; Dodologlu et al., 2003; Silici, 2003).
Undoubtedly, plant extracts and propolis which are considered as alternatives to antibiotics have a wide range of potential uses. Therefore, the determination of the effects of these products on human and animal health are of significant importance at present due to the increasing practice of organic agriculture and increasing importance attached to safe nutrient production. This study, which is inspired by these thoughts, aims to determine the most appropriate utilization levels of propolis and plant extracts either as separate or in combination and their associated effects on animal performance.
MATERIALS AND METHODS
One hundred and five, one-day-old male broiler chicks (Ross 308) were divided into seven treatment groups of 15 birds each and randomly assigned to seven treatment diets. The groups were as follows: 1. Negative control (not including antibiotic), 2. Positive control (including antibiotic), 3. Zingiber officinale (240 ppm), 4. Propolis 1000 ppm, 5. Zingiber officinale 120 ppm+Propolis 500 ppm combination, 6. Zingiber officinale 240 ppm+Propolis 1000 ppm combination, 7. Zingiber officinale 360 ppm+Propolis 1500 ppm combination. The composition of the basal diet is presented in Table 1. Birds were given starter diet from the first day to 10 days, a grower diet from 11 to 21 days, thereafter a finisher diet to 42 days. Each group was fed ad libitum its own diet for a period of 42 days. Twenty four hours light was provided per day. Zingiber officinale essential oils were purchased from Ege Lokman San. Tic. Company, Manisa, Turkey. Propolis (ethanol extracted) was obtained from Erciyes University, S. Cikrikcioglu Junior Technical College-KAYSERI. Major components of Z. officinale essential oil and propolis were analysed using MSGC and are given in Table 2.
The live body weight gains of birds were measured individually and feed consumption and feed conversion efficiency (g feed:g gain) were measured weekly.
Table 1: The ingredient and chemical composition (g/kg) of starter, grower and finisher diets
The efficiency index was calculated by dividing post-experiment live body weight gain (g) by post-experiment feed conversion efficiency. At the end of the experimental period, five birds of similar live body weight from each treatment group were slaughtered to determine blood parameters, carcass weight and dressing percentage, abdominal fat weight and digestive system sections for weight and length. After being stored at refrigerator at +4°C for 24 h, the blood samples were subjected to biochemical (cholesterol, triglycerids and glucose) analysis. CHOD-PAP Method is employed for cholesterol, GPO-PAP Method is employed for triglycerids and Hexokinase Calorimetric-Enzymatic Method was employed for glucose. All analyses were performed by modular DPP device (Roche-Germany). The data obtained in the experiment were subjected to one way analyses of variance to evaluate treatment and random effects using the GLM (General Linear Model) procedure of SAS (1987) in a complete randomized design with seven treatments of 15 replicates. Treatment means were separated using Duncan´s New Multiple Range Test.
Table 2: Zingiber officinale and Propolis essential oil and major components (%)
RESULTS AND DISCUSSION
The data obtained at the experiment regarding cumulative feed consumption, body weight gain, feed conversion efficiency and efficiency index are given in Table 3 on weekly basis. Although treatment effects were not significant (p>0.05), according to Duncan´s test a significant difference (p<0.05) was determined among the groups with respect to feed consumption at the end of the 3rd and 6th week of the experiment. At the end of the experiment, all treatment groups revealed a higher feed consumption pattern (p>0.05) compared to negative control group.
In this study, 240 ppm Z. officinale, 1000 ppm propolis and combination of both had a positive effect on feed consumption with respect to negative control group. This positive effect can be evaluated on the basis of different perspectives. The first of these is the appropriateness of the extract levels for the broilers and associated improvement in the feed taste. The second one is the quicker digestion and passage of the nutrients through the digestive system attributed to the digestive effects of these natural products. Due to this fact, digestive system will have been emptied earlier and feed consumption will have been promoted. These affirmative findings on feed consumption are similar to the previous findings stating that essential oils significantly increase feed consumption in broilers (Alcicek et al., 2004) and that broilers supplemented with propolis had higher feed consumption (Shalmany and Shivazad, 2006).
At the end of the experiment, all treatment groups have experienced higher body weight gains compared to negative control group. These differences were found statistically significant (p<0.05).
Table 3: Effects of Z. officinale essential oil ve propolis supplemental on feed consumption, live body weight gain and feed efficiency of broiler on days 21 and 42
Botsoglou et al. (2002) reported that feed supplemented with 50 and 100 mg kg-1 oregano essential oil did not have any effect on the growth performance of broiler chicks. Similarly, Demir et al. (2003) determined that plant extracts composed of oregano, dusacch, quiponin, garlic and thymol used against antibotic growth promoters did not have any remarkable effect on the body weight gain of broiler chicks. While it was stated that thymol, cinnamaldehyde and CRINA® Poultry, a commercial essential oil mixture, did not affect body weight gain of female broiler chicks (Lee et al., 2003b). Another study proved that antibiotic, probiotic and Genex, a mixture of plant extracts-organic acid, did not have any effect on body weight gain of broiler chicks (Gunal et al., 2006). Considering the studies about the use of propolis in poultry, Biavatti et al. (2003), Sahin et al. (2003), Ziaran et al. (2005) also reported that propolis supplementation did not have any effect on live body weight gain. Contrary to all these studies, there are also studies indicating the positive effects of plant extracts and/or propolis on live body weight gain (Jamroz and Kamel, 2002; Alcicek et al., 2003, 2004; Eclache and Besson, 2004; Hernandez et al., 2004; Sirvydis, 2004; Roodsari et al., 2004; Shalmany and Shivazad, 2006). The findings of our study are also in agreement with those of the mentioned researchers and proved that the two natural products Z. officinale and propolis, significantly improved body weight gain compared to negative control groups. The similar performance of these two natural additives as antibiotic growth promoters is attributed to the decreased number of pathogenic bacteria. Due to the active ingredients in these additives, the formation of a more stable intestinal flora (Tekeli, 2007) and improved feed conversion efficiency in consequence of a better digestion.
At the end of the experiment, no statistically significant differences were determined among the groups with respect to feed conversion efficiency rate (p>0.05). However, when the feed conversion efficiencies of the groups with antibiotic and 240 ppm Z. officinale supplementation were compared with that of the control group, a numerical increase was identified in the former groups. It is supposed that the improvement in feed conversion efficiency is resulted from the increase in appetite due to the stimulation of salivary and gastric glands by Z. officinale extract; the decrease in pathogenic bacteria; formation of a more stable intestinal flora and hence, a better digestibility. Eclache and Besson (2004) revealed that oleo plant extract and avilamycinin had no effect on feed conversion efficiency of broilers. Similarly, there are other findings showing that ration supplemented with plant extract and propolis additives did not have any significant effect on the improvement of feed conversion efficiency of poultry (Demir et al., 2003; Botsoglou et al., 2004; Acikgoz et al., 2004; Ziaran et al., 2005; Gunal et al., 2006). The findings of these researchers are in agreement with those of our study. Roodsari et al. (2004) reported that feed conversion efficiency of broilers has improved along with increased levels of propolis. Quail rations supplemented with 1000 ppm propolis (Denli et al., 2005) and similarly, broiler chick rations supplemented with 200 and 250 ppm propolis alcohol extract (Shalmany and Shivazad, 2006) proved to improve feed conversion efficiency. The unagreement of these findings with those of our study can be attributed to bird material, the amount of propolis used and the different geographic region where it was collected.
With respect to the efficiency index, which is calculated as a component of feed conversion efficiency and live body weight gain; the highest index was recorded in the antibiotic, Z. officinale and 1000 ppm propolis groups, whereas the lowest was recorded in negative control group. These results proved that Z. officinale and propolis additives –though being less effective- performed like anbiotic to certain extent and have a great potential to be utilized as an alternative. The improvement in the performance parameters of 240 ppm Z. officinale and 1000 ppm propolis groups can be explained by the decrease in total erobic mesophilic and coliform bacteria in the jejunum, the increase in lactic acid bacteria and improvement in villi length (Tekeli, 2007).
The findings on the slaughter and carcass characteristics of the birds by the end of the experiment are given in Table 4. Although treatment effects were not significant (p>0.05), according to Duncan´s test a significant differences (p<0.05) were recorded among groups with respect to hot and cold carcass weights. While the highest hot and cold carcass weights were identified in antibotic group, the lowest hot and cold carcass weights were seen in negative control group. Z. officinale and propolis groups performed almost similiar to antibiotics with respect to these parameters. The differences among the groups in carcass weights resulted from the differences in live body weight gains by the end of the experiment. It was identified that different treatments did not have any significant effect on carcass yield (p>0.05). Since there was not any difference among groups with respect to carcass yield, it is considered that feed additives do not have such effect in this regard. Alcicek et al. (2003) reported that an additive of essential oils above 48 ppm (oregano, daphne, sage, myrtle, fennel and citrus oil) did not affect the carcass yield of the broilers.
Table 4: Effects of Z. Officinale essential oil and propolis supplemental on carcass characteristics of broiler chicks
Avci (2004) mentioned that plant extracts of thyme, fennel, ginger, rosemary, nigella and their combination did not have any effect on carcass yield. According to a study of Erener et al. (2005), in broiler chicks, control and carvacrol groups had higher carcass weights compared to menthol group (p<0.05), but no significant difference was determined among groups with respect to carcass yield. In similar manner, Denli et al. (2005) revealed that the propolis supplement in quail rations significantly improved carcass weight compared to control and flavomycin groups while having no effect on carcass yield.
The findings of these researchers are in agreement with the present findings. The difference among the groups with respect to abdominal fat weight and abdominal fat amount % was significant (p<0.05). In this experiment, the lowest abdominal fat weight (19.44 g) was determined in negative control group and the highest abdominal fat weight (32.89 g) was recorded in Z. officinale group. This study showed that the ration additive Z. officinale extract did not decrease, but rather increased the abdominal fat amount compared to the control group. Compared to negative control group, Z. officinale extract generated a significant difference in abdominal fat weight since it possibly promotes fat deposition along with live body weight gain or since the increase in live body weight gain is possibly accompanied by increased fat deposition. It was proved by Erener et al. (2005) that in broilers, carvacrol additive increased abdominal fat weight compared to control and menthol groups. On the other hand, Denli et al. (2005) reported that supplemental propolis in quail rations did not have a significant effect on abdominal fat weight.
It was determined that antibiotic, Z. officinale and propolis extracts had significant effects on some digestion system parameters compared to negative control group (p<0.05) (Table 5).
Table 5: Effect of Z. Officinale essential oil and propolis supplemental on digestive system, liver and heart at 42.days pf broiler shick
While ileum and small intestine weights showed an increase in Z. officinale and propolis supplemented groups, cecum and large intestine weights increased in antibiotic supplemented groups. The longest total digestive system length (202.30 cm) was determined in Z. officinale group and the shortest length (184.48 cm) was observed in negative control group. In similar manner, the highest digestive system weight was determined in treatment groups with respect to negative control group. As also reported by Tekeli et al. (2006, 2008), the additives of antibiotic, Z. officinale and propolis extracts affect the weights and/or lengths of the digestive system. The improvement in such parameters of the digestive system is attributed to the stimulatory and promotive effects of these extracts on the gastric juices and the digestive system. Liver weight was observed to increase in antibiotic and propolis supplemented groups as well as the group supplemented with a 1.5:1.5 combination of propolis and Z. officinale (p<0.05). Alcicek et al. (2004) expressed that herbal essential oil mixtures decreased intestine weights. In a study by Sarica et al. (2005), antibiotic, garlic and thymol when combined with enyzme was reported to lead a significant decrease in small intestine weight and small intestine length significantly increased in control and garlic groups. Cabuk et al. (2006) stated that essential oil mixtures did not have any effect on small intestine weight in broilers. The findings of these researchers are not in agreement with the findings of the subject study. The differences among the findings possibly result from the different plant extracts used, additional enzyme supplementation in the ration and the particular focus of these researchers on a certain portion of the digestive system. Erener et al. (2005) and Cabuk et al. (2006) reported that essential oil mixtures did not have any effect on the weights of either renewable internal organs or pancreas.
The findings on plasma cholesterol, glucose and triglycerids concentrations are presented in Table 6. The difference among groups with respect to plasma cholesterol, glucose and triglycerids values was regarded as significant (p<0.05). The highest plasma cholesterol content (136.40 mg dL-1) was recorded in antibiotic supplemented group. The lowest plasma cholesterol contents, on the other hand, were determined respectively in negative control group (113.60 mg dL-1) and the group supplemented with 240 ppm Z. officinale (118.75 mg dL-1). The findings of the subject study showed that antibiotic increased plasma cholesterol level. This can be explained by the higher live body weight gain induced by antibiotic relative to control group and increased fat deposition due to high calorie intake associated with increased feed consumption.
Table 6: Effect of Z. Officinale essential oil and propolis supplemental on some blood parameters of broiler chicks
While the highest plasma glucose and triglycerids concentrations were identified in control group, the lowest plasma glucose concentrations was observed in the group supplemented with (1.5:1.5) Z. officinale and propolis combination. Similarly, the lowest serum triglycerids concentration was reported in the groups supplemented with (1/2:1/2 and 1:1) Z. officinale and propolis combinations. It was expressed in the findings of diverse studies that plant extract and propolis intake lead to a decrease in the level of plasma cholesterol, glucose and triglycerids concentrations (Lee et al., 2003a; Al-Homidan, 2004; Kaya et al., 2004; Fuliang et al., 2005; El-Bagir et al., 2006). 240 ppm Z. officinale, 500, 1000 ppm propolis and the combinations of these two extracts were particularly effective on lowering plasma triglycerids concentrations. This lowering effect can be attributed to the regulatory mechanism of the flavanoids-as one of the ingredients in these natural products- for blood circulation and stimulation of triglycerids use for energy generation. In the group supplemented with (1.5:1.5) Z. officinale and propolis combination, a decrease in plasma glucose level was noted which possibly resulted from the stimulatory effect of the extracts at this amount on insulin release. It is reported that essential oils are the secondary metabolites of the plants enhancing release of insulin or insulin-like substances (Greathead, 2003). Insulin stimulates glucose transport into liver cells and leads to a decrease in the level of blood glucose. Subsequently, the glucose entered in the liver cells is firstly converted to pyruvate and then to acetyl-CoA, and utilized as substrate in the synthesis of fatty acids. This means increased fat deposition (Guyton and Hall, 2001). Abdominal fat weight was reported to increase in all treatment groups compared to negative control group. Contrary to the findings of this study, Demir et al. (2003), Lee et al. (2003a) and Biavatti et al. (2003) expressed that plant extracts used as growth promoters had no effect on plasma lipid and glucose concentrations. The differences in research findings can possibly be explained by the differences in plant extracts and propolis utilized for the studies as well as the differences in their doses.
Three main remarks can be made on the basis of the findings obtained in this study and their assessment in accordance with the findings of the former studies. The remarks are as follows:
- Extract combinations such as 240 ppm Z. officinale and 1000 ppm propolis, performed similar to antibiotics on live body weight gain, feed consumption and feed conversion efficiency of broiler chicks. The combinations proved superior to antibiotics in terms of lowering plasma cholesterol, glucose and triglycerids levels, and hence, have great potential to replace antibiotic growth promoters
- Z. officinale and propolis –as alternatives to antibiotics- are valuable natural products since they can continuously be used in the ration until slaughter, do not induce any resistance to antibotics, are harmless in apppropriate amounts and are risk-free in terms of residue-formation. The findings are considered important also for the gathering of ecological products
- For a thorough assessment of the studies on the usage of plant extracts and propolis in mixed feed of broiler chicks, it is important to know the location where the extracts are collected, the time of their collection as well as the methods of their extraction. The information on these parameters are crucial in order to further clarify the similarities and differences among the findings of the studies and derive at more precise/undisputed results. Since the effective ingredients in the extracts vary according to the above-mentioned parameters, the active components should be specifically determined through analyses
ACKNOWLEDGMENTS
The authors are grateful to Dr. Sibel Silici from Erciyes University, S. Cikrikcioglu Junior Technical College, Kayseri, for gifts of propolis and to TUBITAK and Cukurova University Research for technical assistance.
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This article was originally published in Science Alert journal in March 25, 2011. Engormix.com thanks the author and the journal for this contribution.