Abstract 

The effect of dietary energy (high and low) and Leucena leaf meal (LLM) levels (at 0 - 20%) on the performance of growing layers was studied using 24 pullets in a digestibility trial and 288 birds aged 12 weeks in a growth study.

The crude protein and crude fibre of LLM were 20.6 and 18.9%, respectively, whereas NFE was only 34.6%. LLM contained about 16.5 MJ/kg DM Gross energy and 1.9% mimosine in DM.  Increasing LLM led to a reduction in most of the parameters measured with the exception of gizzard weights, large intestine length and age at first egg. Plasma glucose concentrations were not affected by treatment, while total plasma proteins fluctuated inconsistently.

This study showed that LLM at 5% had no adverse effect on performance of growing layers but higher inclusion levels affected performance regardless of the dietary energy level. 

Key words: Chicken, energy, growth, leucaena, meal

Introduction 

In many parts of the tropics and subtropics, availability of animal and vegetable protein sources for poultry feeds is unreliable. This has led to a growing need to find locally available substitutes including forages from protein-rich leguminous shrubs such as Leucaena leucocephala and Gliricidia sepium (Leng 1997). These forages are also valuable sources of minerals and vitamins (McDonald et al 1995).  Leucaena leucocephala is a tropical legume cultivated extensively in South East Asia, Latin America, Africa and the West Indies (Lulandala 1985). The leaves, young stems, flowers and pods are excellent sources of protein and minerals. Leaf material of Leucaena compares favourably with Lucerne in terms of crude protein (CP), calcium (Ca) and phosphorus (P), and it is also a good source of b-carotene. Under optimum growing conditions, Leucaena yield large amounts of high quality forage (NAP 1984).

However, despite its nutritive and agronomic advantages, Leucaena contains a tyrosine analogue non-protein amino acid, mimosine (C₈H₁₀N₂O₄), which may be harmful to animals, including poultry (Jones and Megarrity 1983). Leucaena has also very low apparent metabolisable energy which ranges from 2.2 to 2.8 MJ/kg (D'Mello and Acamovic 1982a) associated with the presence of galactomannan gums (Lyon and Kohler 1981).  Since birds eat to satisfy their energy requirements, the inclusion of Leucaena in their feed requires formulations geared at providing sufficient available energy to sustain production and at the same time avoid nutrient wastage and impaired productivity. These factors have not been optimised for poultry. Therefore the present studies were conducted to determine the nutritive value of Leucaena and effects of including Leucaena in diets differing in energy levels, on the performance of growing pullets.

Materials and Methods

Experimental design and dietary composition

Eight treatments were used in two experiments conducted in this study. Leucaena leaf meal (LLM) was added at 0, 5, 10, and 20% of the diets (Table 1). In addition there was optimum (T; 2600-2800 kcal) and high (H; 3000 kcal) dietary energy.  In order to achieve the high energy level, sunflower vegetable oil was added to the H diets. All diets contained equal levels of vitamins and mineral premixes. 

Study I was a digestibility trial conducted for a period of 36 days with the aim of determining the nutritive value of different diets containing varying combinations of LLM and energy in growing pullets. A 4x2 completely randomised factorial design was used involving eight dietary treatments, three experimental periods (week 14, 16 and 18 of age) and three birds per diet. At the end of each week, faeces from one bird were combined, weighed and thoroughly mixed. A small fresh sample was frozen for nitrogen analysis and the remaining sample was dried at 62°C for 48 h, then finely ground and packed in airtight containers for further analyses. During the period of data collection, feed intake was also recorded daily.

Study II was a 12 weeks growth experiment aimed at determining the effect of LLM and dietary energy content on the performance of growing pullets. A complete randomised 4x2 factorial arrangement was used, comprising 8 dietary treatments; each treatment was replicated three times with 12 birds per replicate. The 12 birds in each group were placed in a 120cm x 120cm pen and group fed.  Birds were individually weighed once a week throughout the experimental period. The feed conversion ratio was calculated by dividing the total feed intake by total gain of all the birds in each pen. 

Blood parameters, organs and carcass characteristics 

From week 13 to 22, about 2 ml of blood samples/bird were taken weekly from the brachial vein of four birds in each pen and preserved in Na₂EDTA tubes containing NaF. Plasma samples obtained were stored at –20^oC for glucose and total protein assays.

At 16, 18, 20, and 23 weeks of age, one bird per pen was randomly picked and slaughtered to assess the carcass and organ characteristics. The entire gastro-intestinal tract (GIT) was removed, stretched and measured. The lengths of the oesophagus, duodenum, ileum, caeca and large intestines were expressed as a percentage of the total GIT length. The gizzards were also incised emptied and weighed. Other organs weighed included the heart, liver, ovaries and spleen and were expressed as a percentage of the total carcass weight. Counting the number of growing follicles was used to assess follicular growth.   

Chemical analysis

Analyses of diets and faecal samples were carried out using the proximate method (AOAC 1990) for percentage dry matter (DM), ash, crude protein (CP), ether extract (EE), crude fibre (CF), Ca and P contents. Uric acid nitrogen and faecal nitrogen were measured using a direct chemical method described by Ekman (1954). Ca and P were measured using an ion selective Ion 83 electrode and Hitachi model spectrophotometer (100-20 Japan), respectively.

Plasma glucose and total proteins were measured using respective kits supplied by Randox Laboratories, UK. Mimosine concentration in LLM, diets and faecal samples were measured against the mimosine standard (Sigma) as described by Matsumoto and Sherman (1951). Mimosine was expressed as a percentage of DM. Mimosine load was calculated by the difference between the amount consumed and amount excreted in faeces at different ages. 

Statistical analyses

The treatment effects on various parameters were analysed by the General Linear Model (GLM) procedure of Statistical Analysis Systems (SAS 1988) using a two way Analysis of Variance. Values were only considered to be significant at P<0.05.

Results 

Chemical composition of feed ingredients and experimental diets

The CF content was highest in sunflower meal and lowest in fish meal (Table 2).  LLM had lower CF when compared to sunflower meal. The CP was highest in fish meal and lowest in maize meal but was almost similar in both sunflower seed cake meal and Leucaena leaf meal (LLM). The dry matter content did not vary significantly between feed ingredients (ranging between 95.0% and 89.7%). EE was highest and lowest in maize meal and sunflower meal, respectively. Ca and P levels were highest in fish meal. Sunflower meal had the highest gross energy content. 

Table 3 shows the chemical composition of the compounded treatment diets. The DM content did not differ significantly between the treatment diets The CP content was lowest in the H5 diet, and was almost similar in the other diets. Decreases in CF and EE levels with increasing LLM were observed in all diets. There was no definite trend for ash and NFE in both optimum and high-energy diets. Increase in LLM in diets reduced Ca and P in both the high and optimum energy diets. The gross energy values were highest in T0 and lowest in H10 diets.

Dietary and faecal mimosine

LLM mimosine concentration was 0.4 mg/g (1.9% DM). A significant (P<0.05) increase in mimosine with increasing levels of LLM in the diets was observed (Table 3). The amount of mimosine per g of diet consumed and faeces voided, increased with increasing levels of LLM in both T and H diets at all sampling periods (Table 4). Similarly, in T diets the mimosine load increased as the level of LLM was increased, but changes were inconsistent in birds fed H diets. At 14 weeks of age, dietary treatment did not significantly affect the amount of mimosine excreted. However, increase in LLM in diets led to significant increases in the amount of mimosine excreted in the faeces during the 16^th and 18^th weeks of age.

Digestible nutrients

With the exception of digestible crude protein, digestible nutrients (DM, CF and EE) were found to decrease with increasing levels of LLM (Table 5). 

Effect of dietary treatments on performance

Inclusion of LLM induced yellow coloration of plasma and shanks of the birds. Depth of colour was highest in birds fed the 20% LLM diets and palest in birds fed the 0% LLM diets. The growth rates of birds declined consistently as the LLM levels increased in the diet (Table 6). The highest overall gain was observed in birds fed the optimum energy 0% LLM diet (T 0) and lowest in birds fed the high-energy 20% LLM diet (H 20). 

Overall feed intake decreased with increasing LLM% at both energy levels, the rate of decline being highest in birds given high energy and high (20%) LLM diets. Feed intakes for all the diets were lowest at weeks 16 and 20 of age and thereafter, increased markedly with age up to the 24^th week. Treatment and week interactions were significant (P<0.05). 

Feed required per unit live weight gain (feed conversion) increased  with increasing LLM level in the diets, at both energy levels. However, feed conversion ratio for birds fed 5 and 10% LLM in high-energy diets was similar. Correlation analysis between feed conversion ratio (FCR) and gain showed a negative significant association (r= -0.91, P<0.01) 

Carcass and organ characteristics 

Carcass weight declined with increase in LLM (Table 7). The were no differences in carcass weight between energy levels. The total GIT length, lengths of the oesophagus, duodenum and ileum and weights of the heart and ovaries (expressed as % of carcass weight) increased with age of birds but these changes were not affected by dietary treatment. The liver and gizzard weight were heavier in birds fed 10 and 20 % LLM diets. Significant correlations (P<0.05) were observed between carcass and gizzard weights, while the GIT showed significant (P<0.05) correlations with oesophagus and large intestinal length. Neither treatment nor age of the bird significantly affected caecal and large intestinal lengths. In contrast, there was a significant treatment and age effect on the length of the proventriculus.

Age at first egg was 20, 21, 22, 24, 26 and 29 weeks for birds fed H 0, T 0, H 5, T 5, H 10, T 10 and T 20 diets, respectively and greater than 29 weeks for H 20-fed birds. 

Plasma glucose and total protein concentrations 

There was no significant dietary effect or age effect on plasma glucose levels. Plasma glucose in all birds ranged between 11 and 17.5 mMol.  Mean plasma total protein levels varied between weeks in all groups but the pattern of change was not consistent among treatments.  The plasma total protein levels increased up to week 16, followed by a sharp decline in week 17 and a slight increase in weeks 18 and 19 for all dietary treatments.

Discussion

The proximate and mineral composition of the feed ingredients and LLM used in present studies were within the expected ranges (Mtenga and Laswai 1994; McDonald et al 1995). The mimosine concentration of 1.89% DM compares well with values reported for Leucaena cultivars (D’Mello and Thomas 1978). The low momosine concentration in the compounded diets as compared to levels in the LLM could have been due to the chemical binding of mimosine with other nutrients in the feed. Energy values of 16.49 MJ/kg for LLM observed in the present study were lower than the value of 18.3MJ/kg, reported by Mtenga and Laswai (1994). Yellow shanks and plasma observed in birds fed high LLM diets were probably associated with increased intakes of carotene, present in high amounts (536 mg/kg) in LLM (NAP 1984; Lulandala 1985).

Lower growth rates in birds fed high-energy diets compared to those fed optimum energy diets might be associated with imbalances in ratios between energy and other nutrients, presence of toxic factors and decreased feed intake. The addition of sunflower oil to increase dietary energy might also have contributed to the lower growth rates, since Furuse et al (1992) showed that White Leghorns fed long chain triglycerols had a decreased feed intake and eventually lower body weight gain. 

Progressive increase of LLM in the diets resulted in parallel reductions in growth rates of birds. This was probably due to the antinutritional effects of increased mimosine load and/or poor digestibility of nutrients brought about by the high tannin and fibre fraction of the leaf meal (Arora and Joshi 1986). Additionally, the lowest growth rates observed in birds fed 20% LLM diets, especially those on the high-energy diet, could have been due to low feed intake and the effect of mimosine. Hence, considering the interactive effect of LLM and energy on body weight gain, performance was better at lower LLM levels with high energy, whereas, at optimum energy levels higher LLM levels were favourable.

Feed conversion ratios increased with increases in LLM level in the diet, being highest in birds fed 20% LLM and lowest in birds fed 0% LLM for both optimum and high-energy diets. This could have been due to the effects of mimosine and/or high CF. However, the similar feed conversion values observed in birds fed the 5% LLM and 10% LLM high-energy diets in the present study and other studies (Gloria et al 1966; Reddy et al 1995) indicate that high energy probably alleviates the adverse effects of LLM.

In both optimum and high-energy diets, the carcass weights decreased with increase in LLM, effects being greater in birds fed the 20% LLM high-energy diet. This was probably due to the reduced intake observed in this group, an effect associated with the high fat content of this diet (Isaaks 1963). The observed effects of Leucaena on carcass weights could be an effect traceable to mimosine and the presence of tannin (NAP 1984).

Birds fed the 20% LLM diets had the longest GIT, particularly at week 16 and 18. This could have been a physiological adaptive process to increase surface area required for successful digestion and absorption of the feed containing high crude fibre (Farouq and Odi 1994). However, birds fed either optimum or high-energy diets containing 0% and 5% LLM had longer proventriculus than those fed similar diets at 10% and 20% LLM levels. The cause of this effect was not clear, but could probably be associated with toxic effect of LLM, since mimosine is known to inhibit growth and elongation of the proventriculus as well as its HCl and pepsin activity (Austic and Nesheim 1990).

Increases in liver and gizzard weights with increasing LLM for both energy levels indicate that Leucaena had an effect on these organs. The changes in the liver may be related to the need of this organ to increase its efficiency in detoxifying LLM-derived products, including mimosine (Directo et al 1971) and for the gizzard to handle high fibre diets, resulting from the inclusion of LLM and sunflower meal (Farouq and Odi 1994). 

Birds fed low levels of LLM had low mean age at first egg (20-22 weeks) whereas the lowest ovary weights and highest age at first egg (29 weeks) were observed in birds fed 20% LLM at both energy levels. This concurs with findings reported elsewhere that high dietary LLM levels delay sexual maturity in birds.  The present studies also showed that birds fed different dietary energy and LLM levels varied in age at first egg, indicating that dietary composition has a significant effect on age at sexual maturity in birds (Upase et al 1994; Farouq and Odi 1996).

Dietary treatments induced minimum changes in plasma glucose concentrations, but increasing LLM in diets up to 10% and 20% lowered the plasma protein levels in birds. Plasma glucose levels were within normal levels of 12.79 to 17.36 mMol, as reported by Bell and Freeman (1971). The lowering of plasma protein with increased LLM content in the diets could have been due to reduced intake observed in these birds and/or mimosine induced poor protein utilisation. Increasing mimosine concentrations in faeces (87-99%) with increase in LLM at both energy levels is in agreement with D'Mello and Acamovic (1982^b) who noted about 92% excretion of ingested mimosine by chicks fed LLM diets. Therefore, the deleterious effects observed on feeding LLM-based diets in the present studies may not be wholly associated with mimosine alone. The high dietary crude fibre content might have limited intake and dry matter digestibility, thus causing imbalances in the availability of essential nutrients.

Conclusions 

The findings in the present study show that leucaena leaf meal could be used as a protein substitute at 5% in diets of growing birds. However, further studies to determine long-term effects of leucaena are recommended.

References 

Arora S K and Joshi U N 1986. Lignification of Leucaena leaves during growth and its relationship with mimosine content. Leucaena Research. Report. 7: 34 – 35.

Austic R E and Neishem M C 1990. Nutrient requirements for layers. In: Poultry Production, 13^th Edition. pp 213-221.

AOAC. 1990. Association of Agricultural Chemists Official Methods of Analysis, 15^th Edition, Arlington Virginia, USA pp 807-809.

Bell D J and Freeman B M 1971 Plasma glucose levels, plasma protein levels and sexual maturity in chickens. In: Physiology and Biochemistry of the Domestic Fowl. pp 1483.

Directo C C, Labadan M M and Novilla M N 1971 Clinical pathological responses of SCWL cockerels fed with different levels of ipil-ipil leaf meal. Philippine Journal of Agricultural. Science. 8: 93-105

D’Mello J P F and Thomas D 1978 The nutritive value of dried Leucaena leaf meal from Malawi. Studies with young chicks. Tropical Agriculture 5:45 –50

D’Mello J P F. and Acamovic T 1982a Apparent metabolizable energy value of dried Leucaena leaf meal for young chicks. Tropical Agriculture. 59: 329-332

D’Mello J F P and Acamovic T 1982b Growth performance and mimosine excretion by young chicks fed on Leucaena leucocephala . Animal Feed Science and Technology. 7: 247 – 255

Ekman K 1954. Faeces N determination in poultry manure. Australian Journal of Chemistry. 7: 55.

Farouq G A and Odi H D 1994. Effect of varying calorie and protein levels on layers fed high levels of Leucaena leaf meal. Poultry Science 73: 149-150.

Farouq G A and Odi H D 1996. Long term effect of feeding Leucaena leaf meal on egg production. Poultry Science. 75: Supplement 1: Abstract 146

Furuse M, Mabayo R T, Kita K and Okumura J 1992 Effects of dietary medium chain triglyceride on protein and energy utilisation in growing chicks. British Poultry Science. 33: 49-57.

Gloria L A, GerparcioA L, Aglibut F B and Castillo L S 1966. Leucaena glauca benth for poultry and livestock. III. Protein and energy levels and minerals in minimising toxic effects of mimosine in chick rations. In: Leguminous Leaf Meals in Non-ruminant Nutrition. (Editor: D'Mello J P F). pp 247-277.

Isaaks R E 1963 Fat digestion absorption and transport. In: Physiology and Biochemistry of the Domestic Fowl. (Editors: Bell D J and Freeman B M) 1: 322.

Leng R A 1997  Tree foliage in ruminant nutrition No 139: http://www.fao.org/docrep/003/w7448e/w7448e00.htm 

Lulandala L L 1985 Intercropping Leucaena leucocephala with maize and Beans. PhD Thesis, Sokoine University of Agriculture Morogoro, Tanzania pp 3-6.

Lyon C K and Kohler G O 1981 Leaf Protein concentrates from Leucaena leaves. Leucaena Research Report 2: 81.

Matsumoto H and Sherman G D 1951 Archives of .Biochemistry and Biophysics 33: 195.

McDonald P, Edwards R A and Greenhalgh J F D 1995. Animal Nutrition 5^th Edition. Published by John Wiley and Sons, United States pp 606.

Mtenga L A and Laswai G D 1994 Leucaena leucocephala as feed for rabbits and pigs: detailed chemical composition and effect of level of inclusion on performance. Forest Ecology Management 64: 249-257.

NAP 1984. National Academy Press. Innovations in Tropical Reforestation. In Leucaena: Promising forage and tree crop for the tropics. 2^nd Edition Washington, pp 1-10; 41-51

Reddy S P V V, Ravindra R V, Ahmed N and Sharif S A 1995 Effect of feeding a based diet containing Leucaena leucocephala seeds on chick performance. Nutrition Abstract 65: 4198.

SAS 1988 SAS/STAT User's Guide, Release 6.03 Edition. Cary, NC: SAS Institute Inc., pp 1028.

Upase B T and Jadhav A J 1994. Effect on subabu (Leucaena leucocephala) leaf meal feeding on sexual maturity, feed and economical efficiency of growing layer chicks.  Poultry Advisor 27: 33-36.