Effects of dietary supplement of mulberry leaf (Morus alba) on growth and meat quality in rabbits

Mulberry leaves (Morus alba, MLs) are rich in nutrients, particularly a high protein content and a desirable amino acid composition (Tan et al., 2012; Kumari et al., 2009). Compared with other commonly used feeds, MLs have a higher digestibility and better palatability (Watchanapong et al., 2015). Therefore, the MLs were regarded as a special green forage and strongly recommended by Food and Agricultuer Organisation for use as a protein-rich forage supplement for animal production to replace cereals, e.g. maize, cotton seeds, beans, etc. for saving edible foods in the animal production (Uribe and Sanchez, 2001; Huyen et al., 2012).
       
The utilization of MLs as a nutrient source for rabbits has been studied previously. At the outset, it was advised that MLs could be used as a single feed for adult rabbits (Deshmukh et al., 1993). However, subsequent substitution experiments indicated that the use of MLs in feed should be appropriately limited; otherwise, the body weight gain of the animal would be decreased (Meena et al., 1999; Prasad et al., 2003; Bamikole et al., 2005). It has been reported that the nutritional value and digestibility of mulberry leaves are as good as those of lucerne leaves; however, lucerne leaves can constitute 48% of the fodder in rabbit feed, whereas mulberry leaves are limited to a lower percentage (Premalatha et al., 2012). Bamikole et al., (2005) indicated that half of the rabbit commercial fodder could be replaced by MLs. Prasad et al., (2003) recommended that the proportion of MLs used in rabbit diets should be ≤ 45% and 15% could support a good productive performance. However, which percentage is suitable proportion of MLs in rabbit diet remains un-explored. In this context, the present study was conducted to evaluate the effects of a detailed proportion of mulberry leaf powder (MLP) used as the feed ingredient on growth and slaughter performance, and meat quality in rabbits, to explore an optimum proportion of MLP used in diet and try to analyze the reasons of the limited dosage.

The MLP, processed from green mulberry leaves and new shoots, containing 24.00% crude protein (CP), 24.89% crude fiber, 12.40% moisture and 1.98% fat, was bought from Danyang tianyuanshengshu agriculture science and technology development Co. Ltd. located in Danyang of Jiangsu Province, China. MLP was passed through a 0.25 mm sieve and further processed with other fodders and chemical nutrients into a pellet with the 6.0 mm diameter according to the designed proportion compositions of 0% (CK), 5% (ML5), 10% (ML10), 15% (ML15) and 20% (ML20). All diets were isonitrogenous and isoenergetic. Ingredients and composition of the diets were listed in Table 1. The dry matter (DM) and CP digestibility were evaluated according to an in vitro digestion method (Noblet et al., 2007). The nutrition composition in the compounded fodder meets the national standard of the GB14924.3-2010 issued by National Laboratory Animal Standardization Technical Committee.
 

 
Experimental animals
 
The feeding experiment was conducted from May 9 to June 27 (the first 7 days for adaptation feeding and the subsequent 42 days for measurements), in Qilinshan Soviet Chinchilla Rabbit Farm (Jurong, Jiangsu). Eighty young rabbits (male, 2months), with an initial body weight (IBW) of 1.2~1.4kg, were randomly placed to five groups with sixteen animals in each group. The rabbits were fed in individual cages (45 cm×50 cm×37 cm)made up of cement as the boards with provision of feeder and watering bowls. Feed and water were available ad libitum. To calculate the feed conversion ratio (FCR) and average daily gain (ADG), feed consumption and rabbits body weight (BW) were collected weekly.
 
Slaughter and carcass characteristics
 
At the end of experiment, six rabbits from each group were randomly chosen. The rabbits were weighed, electrically stunned and slaughtered by bleeding of the carotid artery and jugular veins. The bodies were skinned and the head and feet were removed. The carcasses were chilled for 24h at 4°C. Then the carcasses were dissected according to the protocols suggested by Blasco et al., (1993).
 
Meat physical property and chemical composition: The colour of the rabbit longissimus dorsi and hind leg muscles were determined using a Colour Difference Meter (WSC-S, Shanghai Physics and Optics Instrument Co., Shanghai, China). The colours of the meat from perpendicular to the fillet surface at five different locations per sample were evaluated and collected using terms of the L* (lightness), a* (redness) and b* (yellowness) colour space values (Eiben et al., 2010). pH levels were determined according to AOAC (2000). In detail, a 10.0 g sample of the meat muscle was homogenized in 100 mL distilled water, and the mixture was filtered. The pH of the filtrate was measured using a pH meter (Mettler Toledo 320-S, Shanghai Mettler Ltd., China).Water holding capacity(WHC) of the meat was measured according to Hamm (1986) and was expressed as the ratio (×100) of muscle area to total area. Drip losses (after 48 h) in the meat samples were determined as described by Honikel (1998).Moisture, protein, fat and ash content in the longissimus dorsi and hind leg muscles of all animals were determined according to the AOAC (2000) method.
 
Data manipulation and statistical analysis
 
The differences among treatments were statistically analyzed with a one-way ANOVA test in a completely randomized design using Statistical Packages for the Social Sciences (SPSS®, 2001) with the type of diet as the main source of variation. Initial weight was used as a covariate in the statistical model for growth traits. The significant differences among means were compared using Duncan’s new multiple-range test (Duncan, 1955). Data presented as percentages were transformed to the corresponding arcsine values before performing the statistical analysis. P ≤  0.05 was set as the limit of significance.

Effects of MLP on the growth of rabbits
 
As shown in Table 2, ADFI throughout the experimental period showed no significant differences among five treatments (P > 0.05), indicating the nutrient compositions did not change the diet habits of the rabbits. ADG in the ML20 fed rabbits was significantly (P ≤ 0.05) lower than that in the rabbits fed by the three treatments of CK, ML5 and ML10. FCR in the ML20 treatment was significantly (P ≤ 0.05) higher than that in the remaining four treatments including CK. In contrast to the treatment of CK, ADG and FCR were not significantly different among the three treatments of ML5, ML10 and ML15 (P > 0.05).
 

 
Effects of MLP on the carcass characteristics of rabbits: Most of the carcass characteristics (e.g. SW, EW, SEW, DP, SESP, ESP, heart, kidney, spleen and liver) had no significant differences among the five treatments (P > 0.05, Table 3). However, the abdominal fat content showed significant differences between the MLP groups and CK (P ≤ 0.05).
 

 
Effects of MLP on the meat quality of rabbits
 
The items reflecting the meat quality showed a similar trend of changes between the longissimus dorsi and hind leg muscle (Table 4). All the color co-ordinates (L*, a* and b*) in the both longissimus dorsi and hind leg muscles showed significant differences among the MLP groups and CK (P ≤ 0.05). For example, most of the L* and a* values in MLP treatment samples were significantly increased, and the b* value significantly declined than those in the CK (P ≤ 0.05). The drip losses in the ML10, ML15 and ML20 treatments were significantly (P ≤ 0.05) lower than those in CK and moisture, fat and ash values in the ML15 and ML20 treatments were significantly (P ≤ 0.05) lower than those in CK too. However, pH, WHC and protein content in the meat showed no significant differences (P > 0.05) among the five treatments.
 

               
Previous studies demonstrated that MLs helped to improve animal health and growth performance (Liu et al., 2001; Prasad et al., 2003). Our study indicated that MLP for a certain proportion (≤15%) in diet is valuable for improvement of growth for young rabbits, while a high proportion (e.g.20%) had negative effects on the growth performance. Several bioactive compounds in mulberry leaves, including 1-deoxynojirimycin, c-amino butyric acid, polysaccharides and phenols, showed strong antioxidant properties, proved to have strong effects on alleviation of blood sugar and fat resulting in a weight loss (Magdalena et al., 2014; He et al., 2018). Thus, these compounds over feed to animals, particularly at the young stages, may produce negative effects on animal growth through the weight loss, abdominal fat reduction and low fat content in meat. In this study, the average daily feed intake showed no significant differences among five treatments, but a reducing trend in the mulberry treated groups still can be observed. Mulberry has the ability of reducing food intake in a concentration dependent manner by increasing gut transit time (Lee et al., 2008). However, high fibre content might increase the intake of feed in the ML20 group either (Premalatha et al., 2012). The weights of liver and kidney were usually used as the indicators of toxicity in animals because they are detoxification organs (Frederik et al., 2015). In this study the weight of these two organs didn’t differ significantly indicating that there was no serious effect of anti-nutritional factors in the MLs treatments. Considering the analyses above, we suggest that the MLP supplement level ≤ 15% is suitable to rabbits.
       
To our knowledge, the effects of MLP supplement on rabbit meat drip loss and chemical composition have not yet been reported. Our observation that MLP supplement decreased the meat drip loss was in contrast to the conclusion in chicken feed MLP supplement diet (Park and Kim, 2012). This may be in relating to the different animal species, and the accurate effect of MLs on meat drip loss need more advanced determination. The MLP supplement increased the L* and a* values (Table 4), suggesting the rabbit meat became brighter and redder. Previous reports didn’t constitute an agreeable conclusion in this aspect. Martinez et al., (2007) reported that the meat L* and a* values were not remarkable different from that of rabbits feed by Lucerne. Zhou et al., (2014) reported that MLP had no significant effect on the finishing steer meat color. The mechanism of MLP improving rabbit meat appearance is not completely clear, but this may be related to the increase of lycopene and beta-carotene content in meat (Monika, 2012) and also related to changes of the composition, such as moisture, fat and protein in meat and its anti-oxidant properties of constituent bio-active chemicals.

The results showed that MLP for a certain pro portion in diet (≤15%)is greatly valuable for improvement of growth and meat appearance of rabbits and it seriously impede the growth of young rabbits when the proportion up to 20%.