DOI: 10.1111/jpn.12319
ORIGINAL ARTICLE
Effect of different levels of raw and heated grass pea seed (Lathyrus sativus) on nutrient digestibility, intestinal villus morphology and growth performance of broiler chicks A. Riasi1, A. H. Mahdavi1 and E. Bayat2 1 Department of Animal Sciences, College of Agriculture, Isfahan University of Technology, Isfahan, Iran, and 2 Department of Animal Sciences, College of Agriculture, University of Birjand, Birjand, Iran
Summary This study aimed to investigate chemical composition and effect of different levels (0%, 10% and 20%) of raw grass pea (RGP) and heat-treated (120 °C for 30 min) grass pea seed (HGP) on nutrient digestibility, dressing percentage, relative internal organ weights, intestinal villous morphology and broiler chicks’ performance. A total number of 200 day-old male chicks were raised under similar condition for 10 days. On day 11, chicks were randomly assigned to five dietary treatments and four replicates of 10 birds each. The result of chemical analysis indicated that Iranian grass pea seed has low levels of total and condensed tannin, and it may be considered as a good source of protein (36.1%) and energy (17.09 kJ GE/g). Heat treatment reduced (p < 0.05) the total and condensed tannin to 21% and 78% respectively. Grass peas seed had higher levels of nitrogen-free extract, P, Na, Mg and Zn than soya bean meal. The apparent digestibility of gross energy and lipid was affected (p < 0.01) by the treatment diets, and it was the lowest after feeding 20% of HGP (p < 0.05). The relative weight of breast and pancreas (p < 0.05) was affected by treatments. Percentage weight of breast and pancreas increased (p < 0.05) after feeding high levels (20%) of RGP and HGP. Substitution of 20% of RGP and HGP increased the duodenal crypt depth (p < 0.05); however, it had no suppressive effect on villus height as the absorptive surface of intestine. The feed conversion ratio was not affected by the treatments in the total experimental period. This study showed that, although the high level of grass pea seed caused a remarkable increase in the relative weight of pancreas and decreased the apparent digestibility of gross energy and lipid, it had beneficial effect on breast relative weight. It seems that heat processing is not effective method for improving quality of Iranian grass pea seed. Keywords broiler chicks, Lathyrus sativus, processing, growth performance, intestinal morphology Correspondence A. H. Mahdavi, Department of Animal Sciences, College of Agriculture, Isfahan University of Technology, Isfahan 84156-83111, Iran. Tel: +98-31-33913505; Fax: +98-31-33913471; E-mail: [email protected] Received: 12 November 2013; accepted: 3 March 2015
Consumption of poultry products in developing countries has increased over the years, and it has affected the demand for feed and raw materials. It is well known that approximately 90 per cent of total feed cost is used to meet energy and protein requirements of poultry (FAO, 2013). Supplying the traditional energy and protein sources (maize, soya bean meal and fish meal) is costly in developing countries, and for this reason during the two last decades, there has been keen interest for finding and evaluating alternative local feed resources (Chowdhury et al., 2005; Trombetta et al., 2006; FAO, 2013). Grass pea (Lathyrus sativus) is a grain legume adapted to alkaline soils in low and medium rainfall
areas. This plant is quite drought tolerant with no serious disease problem; it is also commonly used in human and animal nutrition (Hanbury and Hughes, 2003). Grass pea seed has a high level of crude protein (CP) (20–32%), and it is superior to other legumes in respect of lysine, calcium and vitamins content (Low et al., 1990; Hanbury et al., 2000). It contains approximately 2700 Kcal ME/kg (Latif and Morris, 1975) with adequate concentrations of most micro- and macrominerals (Low et al., 1990; Rotter et al., 1991). However, grass pea seed contains a variety of antinutritional substances, which may affect its nutritional value for monogastric animals (Hanbury et al., 2000). The most frequently occurring antinutritional substances in this legume are tannins, protease and amylase inhibitors, lectins, non-starch polysaccharides,
Journal of Animal Physiology and Animal Nutrition © 2015 Blackwell Verlag GmbH
1
Introduction
Grass pea seed on broiler performance
phytates and lathyrogens (Riepe et al., 1995; McCann et al., 2006). Therefore, different processing methods such as heating, autoclaving and extruding may be necessary for reduction or inactivation of the antinutritional factors prior to inclusion in poultry feeds. Chowdhury et al. (2005), who studied the nutritional value of grass pea seed for growing and laying pullets, concluded that it may support growth performance of pullets either similar to or better than the control group. They reported that grass pea seed up to 150 g/kg diet was well tolerated for laying hens without any deleterious effects, and there were no symptoms of lathyrism in both grower and laying pullets. Feeding raw or heated grass pea seed up to 367 g/kg diet did not show the typical symptoms of human lathyrism in broiler chicks. However, consumption of raw grass pea seed decreased their growth performance in comparison with the heat-treated seed (Latif and Morris, 1975). Low et al. (1990) and Rotter et al. (1991) indicated that high levels of grass pea seed (600 and 820 g/ kg) in diet of growing broiler chicks had an adverse effect on their growth performance. To our knowledge, there is no information for the effect of grass pea seed on nutrient digestibility, intestinal villous morphology and relative internal organ weights of broiler chicks. These parameters are important for evaluating new protein sources in poultry nutrition and the adverse effects of antinutritional factors. Therefore, the aim of this study was to evaluate the chemical composition of Iranian grass pea seed and the effect of different levels of raw and heat-treated grass pea seed on performance of broiler chicks.
A. Riasi, A. H. Mahdavi and E. Bayat
Samples of raw grass pea seed (RGP) and heat-treated grass pea seed (HGP, 120 °C for 30 min) were grounded to pass through a 1-mm screen and then were subjected to chemical analysis (AOAC, 1995). The concentrations of total and condensed tannins were determined by the method of Makkar and Singh (1992). A total of 200 day-old male chicks (Ross 308) were raised under similar condition for 10 days. On day 11, the chicks were randomly assigned to five dietary treatments with four replicates and 10 birds each as a completely randomized design. The experimental groups received normal isocaloric–isonitrogenous diets substituted with different levels of RGP and HGP (0%, 10% and 20%). Chicks had free access to water and diets during the experimental period. The diets were formulated to meet or exceed nutrient requirements of
chicks during the growing and finishing periods based on Ross broiler manual (Table 1). At the end of each period (days 11–28 and 29–42), weight gain, feed intake and feed conversion ratio (FCR) were determined. The apparent ileal digestibility of gross energy, CP and ether extract (EE) were evaluated using titanium dioxide as an inert marker (Short et al., 1995). Titanium dioxide (0.1% wt/wt) was added to the experimental diets for five consecutive days (day 31–35). On day 35, feed was removed 3 h before slaughtering; three chicks per replicate were randomly selected, anaesthetized and then sacrificed. The Animal Care Advisory Committee of the Isfahan University of Technology approved all experimental procedures and confirmed that the experiments were conducted according to the Iranian Council of Animal Care guidelines (1995). The ileal contents (digesta) were gently squeezed out into separate vials. The excreta and digesta were mixed thoroughly and frozen at 20 °C until further analysis. For evaluation of the intestinal villous morphology, 2-centimetre segments of duodenum (medial portion), jejunum (medial portion posterior to the bile ducts and anterior to Meckel’s diverticulum) and ileum (medial portion posterior to Meckel’s diverticulum and anterior to the ileocecal junction) were collected and immediately fixed in 10% formaldehyde solution (Mahdavi et al., 2010). The histological sections were embedded in paraffin, and transverse and longitudinal sections with 5 lm thickness were prepared using microtome, then stained with haematoxylin–eosin (HE) and examined under the light microscope. At the end of the trial, three chicks were randomly selected and slaughtered for the carcass measurements. Dressing percentage was calculated as percentage of carcass weight to body weight. Weight of legs, breast, liver, pancreas, spleen and abdominal fat were measured and expressed as percentage of body weight. All data were analysed using the GLM procedure of SAS software (SAS Institute, 2001). The model used for data analysis was as follows: Yij = l + Ai + eij, where Yij = observed value for a particular character; l = overall mean; Ai = effect of the ith treatment; and eij = random error associated with the ijth recording. Least squares method was used to identify the significant differences between experimental groups at 0.05 level. Single degree of freedom contrasts were made among treatment means to compare control vs. RGP, control vs. HGP, and also RGP vs. HGP.
2
Journal of Animal Physiology and Animal Nutrition © 2015 Blackwell Verlag GmbH
Material and methods
Grass pea seed on broiler performance
A. Riasi, A. H. Mahdavi and E. Bayat
Table 1 The composition and nutrient content of diets for growing (11–28) and finishing (29–42) periods of broiler chicks Growing*
Finishing*
Ingredient (%)
C
RGP10
RGP20
HGP10
HGP20
C
RGP10
RGP20
HGP10
HGP20
Yellow Corn Soya bean meal Fish Meal RGP HGP† Soya bean Oil Common salt dl-Methionine L-Lysin Dicalcium phosphate Oyster shell Vitamin premix‡ Mineral premix§ Calculated analysis MEn (kJ/g) CP (%) Calcium (%) Available phosphorus (%) Methionine (%) Methionine + cysteine (%) Lysine (%) Tryptophan (%)
63.0 29.1 2.80 – – 1.50 0.20 0.275 0.250 1.40 1.00 0.25 0.25
58.4 24.0 2.80 10.0 – 1.20 0.20 0.300 0.225 1.30 1.075 0.25 0.25
54.2 18.7 2.80 20.0 – 0.80 0.20 0.325 0.200 1.20 1.075 0.25 0.25
58.4 24.0 2.80 – 10.0 1.20 0.20 0.300 0.225 1.30 1.075 0.25 0.25
54.2 18.7 2.80 – 20.0 0.80 0.20 0.325 0.200 1.20 1.075 0.25 0.25
63.6 28.8 1.50 – – 2.60 0.20 0.225 0.150 1.45 0.975 0.25 0.25
59.4 23.5 1.50 10.0 – 2.25 0.20 0.225 0.125 1.35 0.95 0.25 0.25
54.8 18.2 1.50 20.0 – 2.05 0.20 0.275 0.125 1.35 1.025 0.25 0.25
59.4 23.5 1.50 – 10.0 2.25 0.20 0.225 0.125 1.35 0.95 0.25 0.25
54.8 18.2 1.50 – 20.0 2.05 0.20 0.275 0.125 1.35 1.025 0.25 0.25
12.51 21 0.90 0.45 0.47 0.92 1.30 0.22
12.51 21 0.90 0.45 0.47 0.92 1.30 0.22
12.51 21 0.90 0.45 0.47 0.92 1.30 0.22
12.51 21 0.90 0.45 0.47 0.92 1.30 0.22
12.51 21 0.90 0.45 0.47 0.92 1.30 0.22
12.80 18 0.85 0.42 0.40 0.92 1.05 0.18
12.80 18 0.85 0.42 0.40 0.92 1.05 0.18
12.80 18 0.85 0.42 0.40 0.92 1.05 0.18
12.80 18 0.85 0.42 0.40 0.92 1.05 0.18
12.80 18 0.85 0.42 0.40 0.92 1.05 0.18
*Diets with different levels of raw or heat-treated grass pea seed substitution (C: control diet, RGP10 and RGP20: diets with 10% and 20% raw grass pea seed, HGP10 and HGP20: diets with 10% and 20% heated grass pea seed). †HGP was prepared after oven heating of grass pea seed at 120 °C for 30 min. ‡Vitamin premix provided the following per kilogram of diet: vitamin A (from vitamin A acetate), 9800 IU; cholecalciferol, 2100 IU; vitamin E (from dl-atocopheryl acetate), 22 IU; riboflavin, 4.4 mg; nicotinamide, 40 mg; calcium pantothenate, 35 mg; menadione, 1.50 mg; folic acid, 0.80 mg; thiamine, 3 mg; pyridoxine, 10 mg; biotin, 1 mg; choline chloride, 560 mg; and ethoxyquin, 125 mg. §Mineral premix provided the following per kilogram of diet: Mn, 65 mg; Zn, 55 mg; Fe, 50 mg; Cu, 8 mg; I [from Ca (IO3)2H2O], 1.8 mg; and Se, 0.30 mg.
The chemical composition of RGP and HGP is shown in Table 2. Heat treatment (120 °C for 30 min) had no effect on the majority of the chemical composition of grass pea seed. Metabolizable energy (ME)
estimated for grass pea seed is 11.30 kJ/g (Latif and Morris, 1975) that is higher than the ME determined for soya bean meal (9.33 kJ/g; NRC, 1994). Iranian grass pea seed had high level of CP (36.1%), and the protein content was higher than the previous report (24–31%) (Hanbury et al., 2000). Aletor et al. (1994) on selected lines of three Lathyrus species reported that average of CP content of grass pea seed was 32.5%. According to Hanbury et al. (2000) and Chowdhury et al. (2005), the concentration of EE in grass pea seed was relatively low. However, it was higher than EE reported for soya bean meal (0.8%; NRC, 1994). Consistent with Chavan (1998), our samples had more than 50% NFE and it was higher than content of soya bean meal (38.7%, Table 2). Ash content of grass pea seed was approximately 4%, which is in the range of previous reports (Hanbury et al., 2000), and was lower than the value determined for soya bean meal (5%). Grass pea seed had higher levels
Journal of Animal Physiology and Animal Nutrition © 2015 Blackwell Verlag GmbH
3
Results and discussion Nutritional research in poultry has centred on identifying barriers of effective digestion and utilization of nutrients. For this reason, poultry nutritionists have increasingly combined their expertise with that of specialists in other biological sciences, including immunology, histology and molecular biology (FAO, 2013). The grass pea seed, as a grain legume, may have the potential to be used in poultry diet; additionally, recognition of its nutritional facts and some of the histological effects are important. Chemical analysis
Grass pea seed on broiler performance
A. Riasi, A. H. Mahdavi and E. Bayat
Table 2 Chemical composition of raw and heat-treated grass pea seeds compared to soya bean meal (DM basis) Chemical analysis
RGP*
Dry matter (%) Gross energy (kJ/g) Crude protein (%) Ether extract (%) Ash (%) Crude fibre (%) Nitrogen-free extract (NFE) (%)‡ Calcium (%) Available phosphorus (%) Sodium (%) Potassium (%) Chloride (%) Magnesium (%) Copper (mg/kg) Zinc (mg/kg) Total tannin (%) Condensed tannin (%)
95.3 17.09 36.1 1.28 4.22 5.92 53.7 0.06 0.31 0.26 1.42 0.05 0.84 21.3 53.2 0.413 0.177
HGP*
0.20 0.51 1.30 0.80 0.14 0.35 2.10 0.01 0.02 0.01 0.18 0.01 0.21 1.20 2.70 0.05a 0.02a
97.5 17.43 36.1 1.38 4.22 5.84 55.8 0.06 0.31 0.26 1.42 0.05 0.84 20.8 53.7 0.323 0.039
Soya bean meal†
0.30 0.46 1.10 0.12 0.17 0.38 2.30 0.01 0.02 0.01 0.14 0.01 0.24 1.70 2.20 0.07b 0.01b
89.0 – 44.0 0.80 5.50 7.00 38.7 0.29 0.27 0.01 2.00 0.05 0.27 22.0 40.0 – –
a,b Means with no common superscript between RGP and HGP groups are significantly different (P < 0.05). *RGP, raw grass pea seed and HGP, oven-heated grass pea seed at 120 °C for 30 min. All chemical compositions of RGP and HGP were analysed according to AOAC (1995) except non-fibrous carbohydrates. †Adapted from National Research Council (1994). ‡By difference from 100–(moisture + crude protein + lipid + ash + crude fibre).
of P, Na, Mg and Zn, but lower levels of Ca, K and Cu compared to reported data for soya bean meal (NRC, 1994). The concentrations of total and condensed tannins were 0.413% and 0.177% for RGP, and 0.323% and 0.039% for HGP respectively. These data note that heat treatment could reduce the total and condensed tannins of grass pea seed (p < 0.05). One of the most frequently occurring antinutritional substances in the grass pea seed is tannins (Ramachandran and Ray, 2008). Tannins are polyphenolic compounds that classify to the groups of low molecular weight (hydrolysable) and higher molecular weight nonhydrolysable (condensed). It is suggested that the hydrolysable tannins have little effect on the nutrient digestibility, but the condensed form mostly binds to the proteins and forms indigestible complexes (Urga et al., 1995; Wang et al., 2005). Deshpande and Campbell (1992) found that white or cream-coloured seeds of grass pea were associated with low levels of tannins (total and condensed). Ramachandran and Ray (2008) reported that the total tannin content in grass pea seed was 1.3%, and this factor was markedly reduced by different processing methods. They demonstrated that fermentation, extrusion, autoclaving and germination reduced the tannin content by 80.7%, 80.8%, 75.3% and 46.9% respectively. Our results indicated that Iranian grass pea seed has low levels of total and condensed tannin. Therefore, grass pea seed may be 4
considered as a good source of protein and energy in poultry diets. Apparent digestibility of gross energy, protein and lipid
Apparent digestibility of gross energy, protein and lipid of different diets containing 0%, 10% and 20% of RGP and HGP is shown in Table 3. Only few studies have determined the in vitro and in vivo nutrient digestibility of grass pea seed. Hanbury et al. (2000) reported that L. sativus had higher digestibility than Lathyrus cicera. In our study, the treatments affect apparent digestibility of gross energy and lipid (p < 0.01). The diet containing 20% of HGP had the lowest digestibility of gross energy and lipid. This result may be attributed to higher level of feed intake in this group and therefore higher passage rate of digesta. The contrast analysis showed that nutrient digestibility was the same for control, RGP and HGP groups. It could be related to the lower contents of total and condensed tannins in Iranian grass pea seed (Table 2). For this reason, heat processing had no beneficial effect on nutrient digestibility as well. Several factors such as plant type, cultivar, age of plant, stage of development and environmental conditions govern the tannin content in plants. Chavan (1998) reported Journal of Animal Physiology and Animal Nutrition © 2015 Blackwell Verlag GmbH
Grass pea seed on broiler performance
A. Riasi, A. H. Mahdavi and E. Bayat
that Canadian and Indian grass pea seed had 0.11% and 1.54% of condensed tannins respectively. Intestinal histological changes
The histological characteristic of different parts of small intestine is shown in Table 4. Result showed that the treatments affected (p < 0.05) ratio of villus height to crypt depth (VCR) of duodenum and ileum.
Actually, this result was mainly due to the effect of treatment on crypt depth (p < 0.05) instead of villus height. It is well defined that new epithelial cells are produced by the stem cells that reside at the bottom of the crypts and migrate along with the villi to the top (Schat and Myers, 1991). On the other hand, low ratio of villus height to crypt depth may indicate faster tissue turnover, suggesting that a higher demand is required to compensate for normal sloughing or atro-
Table 3 Apparent digestibility of gross energy, protein and lipid in broiler chicks fed different levels of raw and heat-treated grass pea seed (RGP and HGP* respectively) Apparent digestibility (%) Treatments
Gross energy
Protein
Lipid
Control (C) RGP (%) 10 20 HGP (%) 10 20 SEM p-Value Treatment Contrasts C vs. RGP C vs. HGP RGP vs. HGP
91.34a
87.72
91.44ab
92.54a 89.27a
86.75 86.00
92.14ab 91.57b
94.62a 85.13b 1.37
88.35 83.98 1.79
94.71a 85.24c 1.15
0.01
0.54
0.01
0.13 0.70 0.30
0.35 0.23 0.94
0.71 0.12 0.51
Means with no common superscript within each column are significantly (p < 0.05) different. *HGP was prepared after oven heating of grass pea seed at 120 °C for 30 min. a–c
Table 4 Effects of different levels of raw and heat-treated grass pea seeds (RGP and HGP* respectively) on intestinal histological changes of broiler chickens Duodenum
Treatments Control (C) RGP (%) 10 20 HGP (%) 10 20 SEM p-Value Treatments Contrasts C vs. RGP C vs. HGP RGP vs. HGP
Jejunum
Ileum
Villus height (lm)
Crypt depth (lm)
VCR
Villus height (lm)
Crypt depth (lm)
Villus height (lm)
Crypt depth (lm)
VCR
853.7
74.1b
11.52a
557.5
78.3
7.12
352.5
55.0
6.40a
805.8 766.6
63.2b 93.0a
12.75a 8.24b
581.6 606.0
66.0 60.0
8.81 10.10
280.0 291.6
45.0 50.0
6.22ab 5.83ab
767.5 693.3 8.30
60.0b 98.0a 1.45
12.79a 7.07b 1.72
580.0 502.5 4.30
65.0 65.8 1.20
8.92 7.64 2.11
358.3 326.3 4.04
50.0 87.7 1.50
7.17a 3.72b 1.40
0.39
0.04
0.03
0.22
0.88
0.14
0.11
0.43
0.02
0.44 0.06 0.65
0.31 0.94 0.28
0.12 0.14 0.01
0.64 0.95 0.81
0.19 0.11 0.65
0.26 0.20 0.43
0.94 0.79 0.44
0.01 0.21 0.60
0.09 0.53 0.97
VCR
VCR, Villus height: crypt depth ratio. a–b Means with no common superscript within each column are significantly (p < 0.05) different. *HGP was prepared after oven heating of grass pea seed at 120 °C for 30 min.
Journal of Animal Physiology and Animal Nutrition © 2015 Blackwell Verlag GmbH
5
Grass pea seed on broiler performance
A. Riasi, A. H. Mahdavi and E. Bayat
phy of villus due to inflammation from antinutritional factors. Our results showed that feeding 20% of RGP and HGP decreased (p < 0.05) the duodenum VCR. The contrast analysis showed RGP vs. HGP had significant difference (p < 0.01) for duodenum VCR and C vs. RGP had significant difference (p < 0.01) for ileum crypt depth. This finding may be due to the effect of some antinutritional factors (e.g. trypsin inhibitor and non-starch polysaccharides) resistant to heat
processing in grass pea seed (Riepe et al., 1995; McCann et al., 2006). Feng et al. (2007) reported that soya bean meal compared with fermented soya bean meal caused a significant decrease in villus height and a significant increase in crypt depth. They suggested that antinutritional factors in soya bean meal, such as trypsin inhibitor and soya bean globulins had an adverse effect on the morphology and function of digestive tract of birds.
Table 5 Effects of different levels of raw and heat-treated grass pea seeds (RGP and HGP* respectively) on broiler chicks’ performance Feed intake (g/bird) Treatments Control (C) RGP (%) 10 20 HGP (%) 10 20 SEM p-Value Treatment Contrasts C vs. RGP C vs. HGP RGP vs. HGP
11–28 1152.1
Weight gain (g/bird)
29–42 c
1205.7
11–42 b
c
FCR (feed/gain)
11–28
29–42
b
b
1525.6
1.91
2357.8
585.1
941.8
11–42
11–28 a
29–42 1.33
11–42
c
1.58
1269.6a 1258.4b
1339.6a 1334.7a
2636.2a 2593.2b
708.8a 708.9a
950.7a 987.9a
1648.6 1661.5
1.84ab 1.82b
1.46a 1.41b
1.61 1.58
1261.3b 1303.5a 3.69
1354.0a 1333.0a 7.98
2616.2ab 2626.5a 10.12
672.5a 682.6a 16.60
974.6a 922.5a 24.60
1637.4 1607.7 27.10
1.89a 1.90a 0.04
1.41b 1.47a 0.03
1.60 1.64 0.03
0.01
0.01
0.02
0.01
0.04
0.08
0.03
0.01
0.06
ORIGINAL ARTICLE
Effect of different levels of raw and heated grass pea seed (Lathyrus sativus) on nutrient digestibility, intestinal villus morphology and growth performance of broiler chicks A. Riasi1, A. H. Mahdavi1 and E. Bayat2 1 Department of Animal Sciences, College of Agriculture, Isfahan University of Technology, Isfahan, Iran, and 2 Department of Animal Sciences, College of Agriculture, University of Birjand, Birjand, Iran
Summary This study aimed to investigate chemical composition and effect of different levels (0%, 10% and 20%) of raw grass pea (RGP) and heat-treated (120 °C for 30 min) grass pea seed (HGP) on nutrient digestibility, dressing percentage, relative internal organ weights, intestinal villous morphology and broiler chicks’ performance. A total number of 200 day-old male chicks were raised under similar condition for 10 days. On day 11, chicks were randomly assigned to five dietary treatments and four replicates of 10 birds each. The result of chemical analysis indicated that Iranian grass pea seed has low levels of total and condensed tannin, and it may be considered as a good source of protein (36.1%) and energy (17.09 kJ GE/g). Heat treatment reduced (p < 0.05) the total and condensed tannin to 21% and 78% respectively. Grass peas seed had higher levels of nitrogen-free extract, P, Na, Mg and Zn than soya bean meal. The apparent digestibility of gross energy and lipid was affected (p < 0.01) by the treatment diets, and it was the lowest after feeding 20% of HGP (p < 0.05). The relative weight of breast and pancreas (p < 0.05) was affected by treatments. Percentage weight of breast and pancreas increased (p < 0.05) after feeding high levels (20%) of RGP and HGP. Substitution of 20% of RGP and HGP increased the duodenal crypt depth (p < 0.05); however, it had no suppressive effect on villus height as the absorptive surface of intestine. The feed conversion ratio was not affected by the treatments in the total experimental period. This study showed that, although the high level of grass pea seed caused a remarkable increase in the relative weight of pancreas and decreased the apparent digestibility of gross energy and lipid, it had beneficial effect on breast relative weight. It seems that heat processing is not effective method for improving quality of Iranian grass pea seed. Keywords broiler chicks, Lathyrus sativus, processing, growth performance, intestinal morphology Correspondence A. H. Mahdavi, Department of Animal Sciences, College of Agriculture, Isfahan University of Technology, Isfahan 84156-83111, Iran. Tel: +98-31-33913505; Fax: +98-31-33913471; E-mail: [email protected] Received: 12 November 2013; accepted: 3 March 2015
Consumption of poultry products in developing countries has increased over the years, and it has affected the demand for feed and raw materials. It is well known that approximately 90 per cent of total feed cost is used to meet energy and protein requirements of poultry (FAO, 2013). Supplying the traditional energy and protein sources (maize, soya bean meal and fish meal) is costly in developing countries, and for this reason during the two last decades, there has been keen interest for finding and evaluating alternative local feed resources (Chowdhury et al., 2005; Trombetta et al., 2006; FAO, 2013). Grass pea (Lathyrus sativus) is a grain legume adapted to alkaline soils in low and medium rainfall
areas. This plant is quite drought tolerant with no serious disease problem; it is also commonly used in human and animal nutrition (Hanbury and Hughes, 2003). Grass pea seed has a high level of crude protein (CP) (20–32%), and it is superior to other legumes in respect of lysine, calcium and vitamins content (Low et al., 1990; Hanbury et al., 2000). It contains approximately 2700 Kcal ME/kg (Latif and Morris, 1975) with adequate concentrations of most micro- and macrominerals (Low et al., 1990; Rotter et al., 1991). However, grass pea seed contains a variety of antinutritional substances, which may affect its nutritional value for monogastric animals (Hanbury et al., 2000). The most frequently occurring antinutritional substances in this legume are tannins, protease and amylase inhibitors, lectins, non-starch polysaccharides,
Journal of Animal Physiology and Animal Nutrition © 2015 Blackwell Verlag GmbH
1
Introduction
Grass pea seed on broiler performance
phytates and lathyrogens (Riepe et al., 1995; McCann et al., 2006). Therefore, different processing methods such as heating, autoclaving and extruding may be necessary for reduction or inactivation of the antinutritional factors prior to inclusion in poultry feeds. Chowdhury et al. (2005), who studied the nutritional value of grass pea seed for growing and laying pullets, concluded that it may support growth performance of pullets either similar to or better than the control group. They reported that grass pea seed up to 150 g/kg diet was well tolerated for laying hens without any deleterious effects, and there were no symptoms of lathyrism in both grower and laying pullets. Feeding raw or heated grass pea seed up to 367 g/kg diet did not show the typical symptoms of human lathyrism in broiler chicks. However, consumption of raw grass pea seed decreased their growth performance in comparison with the heat-treated seed (Latif and Morris, 1975). Low et al. (1990) and Rotter et al. (1991) indicated that high levels of grass pea seed (600 and 820 g/ kg) in diet of growing broiler chicks had an adverse effect on their growth performance. To our knowledge, there is no information for the effect of grass pea seed on nutrient digestibility, intestinal villous morphology and relative internal organ weights of broiler chicks. These parameters are important for evaluating new protein sources in poultry nutrition and the adverse effects of antinutritional factors. Therefore, the aim of this study was to evaluate the chemical composition of Iranian grass pea seed and the effect of different levels of raw and heat-treated grass pea seed on performance of broiler chicks.
A. Riasi, A. H. Mahdavi and E. Bayat
Samples of raw grass pea seed (RGP) and heat-treated grass pea seed (HGP, 120 °C for 30 min) were grounded to pass through a 1-mm screen and then were subjected to chemical analysis (AOAC, 1995). The concentrations of total and condensed tannins were determined by the method of Makkar and Singh (1992). A total of 200 day-old male chicks (Ross 308) were raised under similar condition for 10 days. On day 11, the chicks were randomly assigned to five dietary treatments with four replicates and 10 birds each as a completely randomized design. The experimental groups received normal isocaloric–isonitrogenous diets substituted with different levels of RGP and HGP (0%, 10% and 20%). Chicks had free access to water and diets during the experimental period. The diets were formulated to meet or exceed nutrient requirements of
chicks during the growing and finishing periods based on Ross broiler manual (Table 1). At the end of each period (days 11–28 and 29–42), weight gain, feed intake and feed conversion ratio (FCR) were determined. The apparent ileal digestibility of gross energy, CP and ether extract (EE) were evaluated using titanium dioxide as an inert marker (Short et al., 1995). Titanium dioxide (0.1% wt/wt) was added to the experimental diets for five consecutive days (day 31–35). On day 35, feed was removed 3 h before slaughtering; three chicks per replicate were randomly selected, anaesthetized and then sacrificed. The Animal Care Advisory Committee of the Isfahan University of Technology approved all experimental procedures and confirmed that the experiments were conducted according to the Iranian Council of Animal Care guidelines (1995). The ileal contents (digesta) were gently squeezed out into separate vials. The excreta and digesta were mixed thoroughly and frozen at 20 °C until further analysis. For evaluation of the intestinal villous morphology, 2-centimetre segments of duodenum (medial portion), jejunum (medial portion posterior to the bile ducts and anterior to Meckel’s diverticulum) and ileum (medial portion posterior to Meckel’s diverticulum and anterior to the ileocecal junction) were collected and immediately fixed in 10% formaldehyde solution (Mahdavi et al., 2010). The histological sections were embedded in paraffin, and transverse and longitudinal sections with 5 lm thickness were prepared using microtome, then stained with haematoxylin–eosin (HE) and examined under the light microscope. At the end of the trial, three chicks were randomly selected and slaughtered for the carcass measurements. Dressing percentage was calculated as percentage of carcass weight to body weight. Weight of legs, breast, liver, pancreas, spleen and abdominal fat were measured and expressed as percentage of body weight. All data were analysed using the GLM procedure of SAS software (SAS Institute, 2001). The model used for data analysis was as follows: Yij = l + Ai + eij, where Yij = observed value for a particular character; l = overall mean; Ai = effect of the ith treatment; and eij = random error associated with the ijth recording. Least squares method was used to identify the significant differences between experimental groups at 0.05 level. Single degree of freedom contrasts were made among treatment means to compare control vs. RGP, control vs. HGP, and also RGP vs. HGP.
2
Journal of Animal Physiology and Animal Nutrition © 2015 Blackwell Verlag GmbH
Material and methods
Grass pea seed on broiler performance
A. Riasi, A. H. Mahdavi and E. Bayat
Table 1 The composition and nutrient content of diets for growing (11–28) and finishing (29–42) periods of broiler chicks Growing*
Finishing*
Ingredient (%)
C
RGP10
RGP20
HGP10
HGP20
C
RGP10
RGP20
HGP10
HGP20
Yellow Corn Soya bean meal Fish Meal RGP HGP† Soya bean Oil Common salt dl-Methionine L-Lysin Dicalcium phosphate Oyster shell Vitamin premix‡ Mineral premix§ Calculated analysis MEn (kJ/g) CP (%) Calcium (%) Available phosphorus (%) Methionine (%) Methionine + cysteine (%) Lysine (%) Tryptophan (%)
63.0 29.1 2.80 – – 1.50 0.20 0.275 0.250 1.40 1.00 0.25 0.25
58.4 24.0 2.80 10.0 – 1.20 0.20 0.300 0.225 1.30 1.075 0.25 0.25
54.2 18.7 2.80 20.0 – 0.80 0.20 0.325 0.200 1.20 1.075 0.25 0.25
58.4 24.0 2.80 – 10.0 1.20 0.20 0.300 0.225 1.30 1.075 0.25 0.25
54.2 18.7 2.80 – 20.0 0.80 0.20 0.325 0.200 1.20 1.075 0.25 0.25
63.6 28.8 1.50 – – 2.60 0.20 0.225 0.150 1.45 0.975 0.25 0.25
59.4 23.5 1.50 10.0 – 2.25 0.20 0.225 0.125 1.35 0.95 0.25 0.25
54.8 18.2 1.50 20.0 – 2.05 0.20 0.275 0.125 1.35 1.025 0.25 0.25
59.4 23.5 1.50 – 10.0 2.25 0.20 0.225 0.125 1.35 0.95 0.25 0.25
54.8 18.2 1.50 – 20.0 2.05 0.20 0.275 0.125 1.35 1.025 0.25 0.25
12.51 21 0.90 0.45 0.47 0.92 1.30 0.22
12.51 21 0.90 0.45 0.47 0.92 1.30 0.22
12.51 21 0.90 0.45 0.47 0.92 1.30 0.22
12.51 21 0.90 0.45 0.47 0.92 1.30 0.22
12.51 21 0.90 0.45 0.47 0.92 1.30 0.22
12.80 18 0.85 0.42 0.40 0.92 1.05 0.18
12.80 18 0.85 0.42 0.40 0.92 1.05 0.18
12.80 18 0.85 0.42 0.40 0.92 1.05 0.18
12.80 18 0.85 0.42 0.40 0.92 1.05 0.18
12.80 18 0.85 0.42 0.40 0.92 1.05 0.18
*Diets with different levels of raw or heat-treated grass pea seed substitution (C: control diet, RGP10 and RGP20: diets with 10% and 20% raw grass pea seed, HGP10 and HGP20: diets with 10% and 20% heated grass pea seed). †HGP was prepared after oven heating of grass pea seed at 120 °C for 30 min. ‡Vitamin premix provided the following per kilogram of diet: vitamin A (from vitamin A acetate), 9800 IU; cholecalciferol, 2100 IU; vitamin E (from dl-atocopheryl acetate), 22 IU; riboflavin, 4.4 mg; nicotinamide, 40 mg; calcium pantothenate, 35 mg; menadione, 1.50 mg; folic acid, 0.80 mg; thiamine, 3 mg; pyridoxine, 10 mg; biotin, 1 mg; choline chloride, 560 mg; and ethoxyquin, 125 mg. §Mineral premix provided the following per kilogram of diet: Mn, 65 mg; Zn, 55 mg; Fe, 50 mg; Cu, 8 mg; I [from Ca (IO3)2H2O], 1.8 mg; and Se, 0.30 mg.
The chemical composition of RGP and HGP is shown in Table 2. Heat treatment (120 °C for 30 min) had no effect on the majority of the chemical composition of grass pea seed. Metabolizable energy (ME)
estimated for grass pea seed is 11.30 kJ/g (Latif and Morris, 1975) that is higher than the ME determined for soya bean meal (9.33 kJ/g; NRC, 1994). Iranian grass pea seed had high level of CP (36.1%), and the protein content was higher than the previous report (24–31%) (Hanbury et al., 2000). Aletor et al. (1994) on selected lines of three Lathyrus species reported that average of CP content of grass pea seed was 32.5%. According to Hanbury et al. (2000) and Chowdhury et al. (2005), the concentration of EE in grass pea seed was relatively low. However, it was higher than EE reported for soya bean meal (0.8%; NRC, 1994). Consistent with Chavan (1998), our samples had more than 50% NFE and it was higher than content of soya bean meal (38.7%, Table 2). Ash content of grass pea seed was approximately 4%, which is in the range of previous reports (Hanbury et al., 2000), and was lower than the value determined for soya bean meal (5%). Grass pea seed had higher levels
Journal of Animal Physiology and Animal Nutrition © 2015 Blackwell Verlag GmbH
3
Results and discussion Nutritional research in poultry has centred on identifying barriers of effective digestion and utilization of nutrients. For this reason, poultry nutritionists have increasingly combined their expertise with that of specialists in other biological sciences, including immunology, histology and molecular biology (FAO, 2013). The grass pea seed, as a grain legume, may have the potential to be used in poultry diet; additionally, recognition of its nutritional facts and some of the histological effects are important. Chemical analysis
Grass pea seed on broiler performance
A. Riasi, A. H. Mahdavi and E. Bayat
Table 2 Chemical composition of raw and heat-treated grass pea seeds compared to soya bean meal (DM basis) Chemical analysis
RGP*
Dry matter (%) Gross energy (kJ/g) Crude protein (%) Ether extract (%) Ash (%) Crude fibre (%) Nitrogen-free extract (NFE) (%)‡ Calcium (%) Available phosphorus (%) Sodium (%) Potassium (%) Chloride (%) Magnesium (%) Copper (mg/kg) Zinc (mg/kg) Total tannin (%) Condensed tannin (%)
95.3 17.09 36.1 1.28 4.22 5.92 53.7 0.06 0.31 0.26 1.42 0.05 0.84 21.3 53.2 0.413 0.177
HGP*
0.20 0.51 1.30 0.80 0.14 0.35 2.10 0.01 0.02 0.01 0.18 0.01 0.21 1.20 2.70 0.05a 0.02a
97.5 17.43 36.1 1.38 4.22 5.84 55.8 0.06 0.31 0.26 1.42 0.05 0.84 20.8 53.7 0.323 0.039
Soya bean meal†
0.30 0.46 1.10 0.12 0.17 0.38 2.30 0.01 0.02 0.01 0.14 0.01 0.24 1.70 2.20 0.07b 0.01b
89.0 – 44.0 0.80 5.50 7.00 38.7 0.29 0.27 0.01 2.00 0.05 0.27 22.0 40.0 – –
a,b Means with no common superscript between RGP and HGP groups are significantly different (P < 0.05). *RGP, raw grass pea seed and HGP, oven-heated grass pea seed at 120 °C for 30 min. All chemical compositions of RGP and HGP were analysed according to AOAC (1995) except non-fibrous carbohydrates. †Adapted from National Research Council (1994). ‡By difference from 100–(moisture + crude protein + lipid + ash + crude fibre).
of P, Na, Mg and Zn, but lower levels of Ca, K and Cu compared to reported data for soya bean meal (NRC, 1994). The concentrations of total and condensed tannins were 0.413% and 0.177% for RGP, and 0.323% and 0.039% for HGP respectively. These data note that heat treatment could reduce the total and condensed tannins of grass pea seed (p < 0.05). One of the most frequently occurring antinutritional substances in the grass pea seed is tannins (Ramachandran and Ray, 2008). Tannins are polyphenolic compounds that classify to the groups of low molecular weight (hydrolysable) and higher molecular weight nonhydrolysable (condensed). It is suggested that the hydrolysable tannins have little effect on the nutrient digestibility, but the condensed form mostly binds to the proteins and forms indigestible complexes (Urga et al., 1995; Wang et al., 2005). Deshpande and Campbell (1992) found that white or cream-coloured seeds of grass pea were associated with low levels of tannins (total and condensed). Ramachandran and Ray (2008) reported that the total tannin content in grass pea seed was 1.3%, and this factor was markedly reduced by different processing methods. They demonstrated that fermentation, extrusion, autoclaving and germination reduced the tannin content by 80.7%, 80.8%, 75.3% and 46.9% respectively. Our results indicated that Iranian grass pea seed has low levels of total and condensed tannin. Therefore, grass pea seed may be 4
considered as a good source of protein and energy in poultry diets. Apparent digestibility of gross energy, protein and lipid
Apparent digestibility of gross energy, protein and lipid of different diets containing 0%, 10% and 20% of RGP and HGP is shown in Table 3. Only few studies have determined the in vitro and in vivo nutrient digestibility of grass pea seed. Hanbury et al. (2000) reported that L. sativus had higher digestibility than Lathyrus cicera. In our study, the treatments affect apparent digestibility of gross energy and lipid (p < 0.01). The diet containing 20% of HGP had the lowest digestibility of gross energy and lipid. This result may be attributed to higher level of feed intake in this group and therefore higher passage rate of digesta. The contrast analysis showed that nutrient digestibility was the same for control, RGP and HGP groups. It could be related to the lower contents of total and condensed tannins in Iranian grass pea seed (Table 2). For this reason, heat processing had no beneficial effect on nutrient digestibility as well. Several factors such as plant type, cultivar, age of plant, stage of development and environmental conditions govern the tannin content in plants. Chavan (1998) reported Journal of Animal Physiology and Animal Nutrition © 2015 Blackwell Verlag GmbH
Grass pea seed on broiler performance
A. Riasi, A. H. Mahdavi and E. Bayat
that Canadian and Indian grass pea seed had 0.11% and 1.54% of condensed tannins respectively. Intestinal histological changes
The histological characteristic of different parts of small intestine is shown in Table 4. Result showed that the treatments affected (p < 0.05) ratio of villus height to crypt depth (VCR) of duodenum and ileum.
Actually, this result was mainly due to the effect of treatment on crypt depth (p < 0.05) instead of villus height. It is well defined that new epithelial cells are produced by the stem cells that reside at the bottom of the crypts and migrate along with the villi to the top (Schat and Myers, 1991). On the other hand, low ratio of villus height to crypt depth may indicate faster tissue turnover, suggesting that a higher demand is required to compensate for normal sloughing or atro-
Table 3 Apparent digestibility of gross energy, protein and lipid in broiler chicks fed different levels of raw and heat-treated grass pea seed (RGP and HGP* respectively) Apparent digestibility (%) Treatments
Gross energy
Protein
Lipid
Control (C) RGP (%) 10 20 HGP (%) 10 20 SEM p-Value Treatment Contrasts C vs. RGP C vs. HGP RGP vs. HGP
91.34a
87.72
91.44ab
92.54a 89.27a
86.75 86.00
92.14ab 91.57b
94.62a 85.13b 1.37
88.35 83.98 1.79
94.71a 85.24c 1.15
0.01
0.54
0.01
0.13 0.70 0.30
0.35 0.23 0.94
0.71 0.12 0.51
Means with no common superscript within each column are significantly (p < 0.05) different. *HGP was prepared after oven heating of grass pea seed at 120 °C for 30 min. a–c
Table 4 Effects of different levels of raw and heat-treated grass pea seeds (RGP and HGP* respectively) on intestinal histological changes of broiler chickens Duodenum
Treatments Control (C) RGP (%) 10 20 HGP (%) 10 20 SEM p-Value Treatments Contrasts C vs. RGP C vs. HGP RGP vs. HGP
Jejunum
Ileum
Villus height (lm)
Crypt depth (lm)
VCR
Villus height (lm)
Crypt depth (lm)
Villus height (lm)
Crypt depth (lm)
VCR
853.7
74.1b
11.52a
557.5
78.3
7.12
352.5
55.0
6.40a
805.8 766.6
63.2b 93.0a
12.75a 8.24b
581.6 606.0
66.0 60.0
8.81 10.10
280.0 291.6
45.0 50.0
6.22ab 5.83ab
767.5 693.3 8.30
60.0b 98.0a 1.45
12.79a 7.07b 1.72
580.0 502.5 4.30
65.0 65.8 1.20
8.92 7.64 2.11
358.3 326.3 4.04
50.0 87.7 1.50
7.17a 3.72b 1.40
0.39
0.04
0.03
0.22
0.88
0.14
0.11
0.43
0.02
0.44 0.06 0.65
0.31 0.94 0.28
0.12 0.14 0.01
0.64 0.95 0.81
0.19 0.11 0.65
0.26 0.20 0.43
0.94 0.79 0.44
0.01 0.21 0.60
0.09 0.53 0.97
VCR
VCR, Villus height: crypt depth ratio. a–b Means with no common superscript within each column are significantly (p < 0.05) different. *HGP was prepared after oven heating of grass pea seed at 120 °C for 30 min.
Journal of Animal Physiology and Animal Nutrition © 2015 Blackwell Verlag GmbH
5
Grass pea seed on broiler performance
A. Riasi, A. H. Mahdavi and E. Bayat
phy of villus due to inflammation from antinutritional factors. Our results showed that feeding 20% of RGP and HGP decreased (p < 0.05) the duodenum VCR. The contrast analysis showed RGP vs. HGP had significant difference (p < 0.01) for duodenum VCR and C vs. RGP had significant difference (p < 0.01) for ileum crypt depth. This finding may be due to the effect of some antinutritional factors (e.g. trypsin inhibitor and non-starch polysaccharides) resistant to heat
processing in grass pea seed (Riepe et al., 1995; McCann et al., 2006). Feng et al. (2007) reported that soya bean meal compared with fermented soya bean meal caused a significant decrease in villus height and a significant increase in crypt depth. They suggested that antinutritional factors in soya bean meal, such as trypsin inhibitor and soya bean globulins had an adverse effect on the morphology and function of digestive tract of birds.
Table 5 Effects of different levels of raw and heat-treated grass pea seeds (RGP and HGP* respectively) on broiler chicks’ performance Feed intake (g/bird) Treatments Control (C) RGP (%) 10 20 HGP (%) 10 20 SEM p-Value Treatment Contrasts C vs. RGP C vs. HGP RGP vs. HGP
11–28 1152.1
Weight gain (g/bird)
29–42 c
1205.7
11–42 b
c
FCR (feed/gain)
11–28
29–42
b
b
1525.6
1.91
2357.8
585.1
941.8
11–42
11–28 a
29–42 1.33
11–42
c
1.58
1269.6a 1258.4b
1339.6a 1334.7a
2636.2a 2593.2b
708.8a 708.9a
950.7a 987.9a
1648.6 1661.5
1.84ab 1.82b
1.46a 1.41b
1.61 1.58
1261.3b 1303.5a 3.69
1354.0a 1333.0a 7.98
2616.2ab 2626.5a 10.12
672.5a 682.6a 16.60
974.6a 922.5a 24.60
1637.4 1607.7 27.10
1.89a 1.90a 0.04
1.41b 1.47a 0.03
1.60 1.64 0.03
0.01
0.01
0.02
0.01
0.04
0.08
0.03
0.01
0.06
