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Nutrient composition, polyphenolic contents, and in situ protein degradation kinetics of leaves from three mulberry species
Livestock Science 146 (2012) 203–206
Contents lists available at SciVerse ScienceDirect
Livestock Science journal homepage: www.elsevier.com/locate/livsci
Short communication
Nutrient composition, polyphenolic contents, and in situ protein degradation kinetics of leaves from three mulberry species W.X. Wang, H.J. Yang n, Y.K. Bo, S. Ding, B.H. Cao State Key Laboratory of Animal Nutrition, College of Animal Science and Technology, China Agricultural University (CAU), Beijing 100193, PR China
a r t i c l e i n f o
abstract
Article history: Received 13 February 2012 Received in revised form 12 March 2012 Accepted 19 March 2012
Mulberry leaves of 14 cultivars of Morus atropurpurea Roxb (MAR), 7 cultivars of Morus alba Linn (MAL) and 8 cultivars of Morus multicaulis Perr (MMP) were assessed for their differences in chemical composition and in situ ruminal degradation. The contents of acid detergent fibre (P¼ 0.073) and ether extract (Po0.0001) ranked as: MMP4MAL4MAR. Crude protein (CP), neutral detergent fibre and polyphenolic contents of the leaves did not differ between three species. Nylon bags containing the leaves were incubated for 2, 6, 12, 24, 36, 48 and 72 h in three rumen cannulated dairy cows. Soluble CP fraction and fractional degradation rate did not differ between three species, but the potential degradable CP fraction in the leaves was greater for MAR than the others (P¼ 0.005). Effective CP degradability was consistently ranked: MAR4MAL4MMP (P¼ 0.051). Nontannin phenols and total tannins were negatively correlated with the soluble CP fraction (r¼ 0.48, Po0.01) and the potential degradable CP fraction (r¼ 0.36, Po0.05), respectively. In summary, the mulberry leaves with high protein, low fibre and tannin contents could be exceptional forage for use in ruminant animals. & 2012 Elsevier B.V. All rights reserved.
Keywords: Mulberry leaves Polyphenolic content Rumen degradation
1. Introduction Mulberry trees grow under varied climatic conditions, ranging from temperate to tropical in the world. Mulberry leaves are highly palatable and rich in protein (Kandylis et al., 2008), but they also have some anti-nutritional factors, such as tannins (Makkar, 2003). Chemical composition and biomass yield of mulberry leaves vary due to the variety, maturity degree, leaf position within the branch, and fertilisation level (Sanchez, 2000). There are at least 15 species of mulberry known in China, but only the leaves of a few species have been analysed for their phytochemical profiles instead of their nutritive values (Han, 2006). In the present study, mulberry leaves from
Abbreviations: ADF, Acid detergent fibre; CP, Crude protein; DM, Dry matter; NDF, Neutral detergent fibre n Corresponding author. Tel.: þ86 10 6273 3124; fax: þ 86 10 6273 4859. E-mail address: [email protected] (H.J. Yang). 1871-1413/$ - see front matter & 2012 Elsevier B.V. All rights reserved. http://dx.doi.org/10.1016/j.livsci.2012.03.009
29 cultivars from three species, grown under the same climatic and geographic conditions, were harvested in Guangdong province. Since no studies have yet examined for digestibility values of these leaves, the objective of the present study was firstly to assess the nutritive value of these leaves in terms of chemical composition and in situ ruminal protein degradation. Secondly, we attempted to explore the relationships between the polyphenolic contents and the in situ degradation.
2. Materials and methods 2.1. Sample collection Mulberry leaves from 14 cultivars of Morus atropurpurea Roxb (MAR), 7 cultivars of M. alba Linn (MAL) and 8 cultivars of M. multicaulis Perr (MMP), were gathered during the summer season of May and June at the mulberry tree experimental plantation field of South Agricultural University
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W.X. Wang et al. / Livestock Science 146 (2012) 203–206
of China (Guangzhou, Guangdong, China) with an average altitude of 15 m above sea level. Average annual temperature was 21.9 1C, with an average minimum temperature of 13 1C in January, and a maximum medium temperature of 28.5 1C in July. Average annual precipitation was 1800 mm. The fresh leaf samples (400 g) of 5 trees per cultivar were randomly picked at the field site from the canopy at 1.0 m above the ground. The leaf samples from the same cultivar of mulberry trees were pooled and mixed to create 2.0 kg leaf samples. The samples were dried at 65 1C for 48 h and then ground to pass through a 1.18-mm screen for chemical analysis, and a 2 mm screen for in situ rumen incubation. 2.2. Animals and diet Three lactating multiparous Holstein cows, fitted with ruminal cannulae (Type 2c; Bar Diamond Inc., Parma, Idaho, USA), served as experimental animals for in situ rumen incubation. The cows had free access to water and were fed 25 kg total mixed ration divided into two equal meals at 07:00 and 17:00. The ration contained (per kg of dry matter (DM)): 250 g maize silage, 167 g ryegrass hay, 83 g alfalfa hay, 267 g maize meal, 69 g wheat bran, 138 g soybean meal, 11 g limestone, 5.5 g dicalcium phosphate, 4.5 g sodium chloride and 5 g commercial vitamin and trace mineral premix. Net energy for lactation and crude protein (CP) content was 6.69 MJ and 160 g, respectively, per kg DM. 2.3. Rumen incubations Throughout the experimental period, nylon bags (12 8 cm2 i.d.; 48 mm of pore size) were filled with 3 g leaf samples. The bags containing the leaves were sealed using rubber bands, bound to plastic strings and soaked in tap water for 20 min prior to ruminal incubation. Each cultivar leaf was incubated in each cow for the testing incubation time period: 2, 6, 12, 24, 36, 48 and 72 h after the morning feeding. After removal from the incubation, the bags were washed with tap water for 15 min until the washing ran clear and colourless. The bags were dried at 65 1C for 48 h, weighed and then the residues in the bags were used for analysis of CP content. Crude protein disappearance was calculated as the difference between the original incubated CP and the residual CP. Rumen degradation kinetics for CP disappearance per mulberry leaf per cow was calculated using the nonlinear model proposed by Ørskov and McDonald (1979). 2.4. Chemical analysis Representative leaf samples were analysed for content of DM (method 930.5), CP (method 984.13), ether extract (method 920.30), neutral detergent fibre (NDF, method 2002.04, without heat stable amylase), acid detergent fibre (ADF, method 973.18) and ash (ID 942.05) with AOAC methods (2000). Both NDF and ADF were corrected for residual ash content. Leaf samples (200 mg) were weighed into 10 ml 0.70 (v/v) aqueous acetone solution, and then were extracted with ultrasonicator at 4 1C, centrifuged at 3000 g for 20 min. Total phenolic contents in the supernatants were determined
at 725 nm by Folin-Ciocalteau reagent using tannic acid (Sigma Aldrich, St. Louis, USA) as a standard. Total tannin content was measured as the difference between total phenols before and after tannin removal by adsorption on insoluable polyvinylpyrrolidone. Non-tannin phenols content in the leaves was calculated by subtracting total tannins from total phenols content. 2.5. Statistical analyses Chemical composition measurements per mulberry cultivar were averaged before the statistical analysis. A total of 29 mulberry cultivar averages, grouped into MAR (n¼ 14), MAL (n¼7), and MMP (n¼8), were used, and statistical analyses were performed using the general linear model (SAS, 1999): Yij ¼ m þMi þ ej where Y is the dependent variable, m is the overall mean, Mi is the fixed effect of mulberry specie (i¼MAR, MAL, MMP), and ej is the random residual error. In situ degradation of the leaves with three animal replicates resulted in a total of 87 observations. After the observations per mulberry cultivar was averaged within three animals, a total of 29 mulberry cultivar averages, grouped into MAR (n¼14), MAL (n¼7), and MMP (n¼8), were used for the statistical analysis with the same model as above. The means were compared using a multiple comparison test (Tukey). The relationships between polyphenolic contents and in situ degradation kinetics were examined by a correlation analysis (SAS, 1999). Significance was declared at Po0.05 unless significance P values between 0.05 and 0.10 are considered as trends and reported. 3. Results and discussion Mulberry leaf are of a high protein quality comparable to or better than soybean meal (Machii, 1989), and it could be an alternative supplemental feed, comparable and in some cases superior to alfalfa, to replace conventional feed resources for goat (Nguyen et al., 2005) and sheep (Doran et al., 2007). In the present study, CP and NDF contents did not differ between three species (Table 1). Ether extract (Po0.0001) and ADF (P¼0.073) contents ranked as: MMP4 MAL4MAR. The CP content was close to the upper value noted in the literature (180–250 g/kg DM; Fotadar and Dandin, 1997; Omar et al., 1999). Feeds with similar CP availability in the rumen could be used to synchronise nutrient supply for maximum microbial protein yield (Herrera-Saldana et al., 1990). In the present study, the highest potential degradable CP fraction occurred in MAR (P¼0.005), and no differences for soluble CP fraction and fractional degradation rate were observed in the leaves between three species (Table 1). Effective CP degradability ranked as: MAR4MAL4MMP (Po0.051). Compared to the results reported by Woods et al. (2003), the soluble CP fraction in the present study was lower than most protein feeds (e.g., soybean meal and maize gluten); the potential degradable CP fraction was higher; the rate was similar or somewhat higher, and the effective degradability was similar when the passage rate k was assumed as 0.08/h. The degradation rate and effective CP degradability in the present study were quite close to those of MAL leaves observed by Singh and Makkar (2000), who noted that the
W.X. Wang et al. / Livestock Science 146 (2012) 203–206
205
Table 1 Chemical composition and ruminal crude protein degradation kinetics of leaves from Morus atropurpurea Roxb (MAR), Morus alba Linn (MAL) and Morus multicaulis Perr (MMP). MAR
MAL
MMP
S.E.M
P value
Chemical composition (g/kg dry matter) Crude protein Ether extract Neutral detergent fibre Acid detergent fibre Total phenolics Total tannins Non-tannin phenols
251.7a 28.5c 278.8a 164.9b 15.5a 1.2a 14.3a
259.0a 42.1b 262.5a 170.7ab 14.3a 1.3a 13.0a
251.8a 49.1a 275.4a 176.6a 15.2a 1.6a 13.6a
4.03 1.84 5.89 3.00 0.58 0.20 0.44
0.531 o0.0001 0.279 0.073 0.502 0.495 0.239
Rumen degradation kinetics Soluble protein Potential degradable protein Fractional degradation rate Effective degradability
0.06a 0.94a 0.048a 0.43a
0.09a 0.90b 0.046a 0.40b
0.07a 0.89b 0.043a 0.36c
0.014 0.010 0.0027 0.012
0.323 0.005 0.341 0.051
In the table, rumen degradation kinetics were calculated using the nonlinear model proposed by Ørskov and McDonald (1979): y¼aþb (1 e c t) where y¼ degradability for crude protein at time t relative to incubation (h), a¼ soluble fraction, b¼ potential degradable fraction, and c¼fractional degradation rate. Effective degradability was calculated using the equation: ED¼ aþ (bþc)/(cþk) with an assumed passage rate (k) of 0.08/h (Tamminga et al., 1990). Means in a row without a common superscript differ as noted P value.
kinetic parameters of CP in their mulberry leaves were comparable to those of other good quality tree fodders such as Leucaena leucocephala and Ficus roxburghii. As reviewed by McSweeney et al. (2001), tannins are polyphenolic compounds of plant origin, which are of two distinct types, hydrolysable tannins (polyesters of gallic acid and various individual sugars) and condensed tannin (polymers of flavonoids). Condensed tannin content ( o27 mg/kg DM) measured in the present study may be negligible for the mulberry leaves and was not presented in Table 1. No distinct differences occurred for total phenols, total tannins, non-tannin phenols in the leaves between three species (Table 1) in which the total phenols contents in the leaves were slightly less than 18 g/kg DM reported by Singh and Makkar (2000). The presence of tannins in nutritionally important forage trees, shrubs, legumes, cereals and grain often limits their feed utilisation, as evidenced by Silanikove et al. (1997). The tannin compounds can have toxic and/or anti-nutritional effects on animals due to the complexing (tanning) ability with dietary protein, which then inhibits the growth of microorganisms (McSweeney et al., 2001). Non-tannin phenols and total tannins in Table 2 were negatively correlated with the soluble CP fraction (r¼ 0.48, Po0.01) and the potential degradable CP fraction (r¼ 0.36, Po0.05), respectively. These findings were consistent with Liener’s study (1980), in which tannins caused an adverse effect on protein digestibility.
Table 2 Partial correlation coefficients between polyphenolic contents in mulberry leaves (n¼ 29 cultivars) and in situ crude protein degradation kinetics. Total phenol
Total tannins
Non-tannin phenols
Soluble protein 0.42n Potential degradable protein 0.19 Fractional degradation rate 0.36n
0.12 0.36n 0.37n
0.48nn 0.02 0.24
P o0.01. P o 0.05.
nn n
Conflict of interest statement We respectfully submit our manuscript entitled ‘‘Nutrient composition, polyphenolic contents, and in situ protein degradation kinetics of leaves from three mulberry species’’ for your consideration for publication in the Livestock Science, Section of Ruminant Nutrition. All authors agree to this submission and affirm that the manuscript presents original work by the authors and is not being considered by any other journal. Hereby, we want to state that there are no any conflict of interests for the submission of this manuscript.
Appendix A. Supplementary material 4. Conclusion The mulberry leaves appear to have a high potential as a protein-rich forage with low fibre content and could serve as comparatively less degradable protein in the rumen. Although there may be associations between polyphenolic compound deposition and degradability measures that cannot be accounted for through simple correlation analysis, mulberry leaves with very low tannin content could be exceptional forage for ruminant animals.
Supplementary data associated with this article can be found in the online version at doi:10.1016/j.livsci.2012. 03.009.
References AOAC, 2000. Official methods of analysis, 17th ed. Association of Official Analytical Chemists, Arlington, VA, USA.
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Doran, M.P., Laca, E.A., Sainz, R.D., 2007. Total tract and rumen digestibility of mulberry foliage (Morus alba), alfalfa hay and oat hay in sheep. Anim. Feed Sci. Technol. 138, 239–253. Fotadar, R.K., Dandin, S.B., 1997. Chemical composition and feeding studies of different elite mulberry genotypes under temperate conditions. Indian J. Seric. 36, 22–26. Han, S.Y., 2006. Mulberry resources and its diverse utilization. Guizhou Agric. Sci. 34, 118–120. (In Chinese with English abstract). Herrera-Saldana, R., Gomez-Alarcon, R., Torabi, M., Huber, J.T., 1990. Influence of synchronizing protein and starch degradation in the rumen on nutrient utilization and microbial synthesis. J. Dairy Sci. 73, 142–148. Kandylis, K., Hadjigeorgiou, I., Harizanis, P., 2008. The nutritive value of mulberry leaves (Morus alba) as a feed supplement for sheep. Trop. Anim. Health Prod. 41, 17–24. Liener, I.E., 1980. Heat-labile antinutritional factors. In: Summerfield, R.J., Bunting, A.H. (Eds.), Advances in Legume Science, Royal Botanic Gardens, Kew, Richmond, Surrey, UK, pp. 157–170. Machii, H., 1989. Varietal differences of nitrogen and amino acid contents in mulberry leaves. Acta Sericologica Entomol. (Japan) 1, 51–61. Makkar, H.P.S., 2003. Effects and fate of tannins in ruminant animals, adaptation to tannins, and strategies to overcome detrimental effects of feeding tannin-rich feeds. Small Ruminant Res. 49, 241–256. McSweeney, C.S., Palmer, B., McNeill, D.M., Krause, D.O., 2001. Microbial interaction with tannins: nutritional consequences for ruminant. Anim. Feed Sci. Technol. 91, 83–93. Nguyen, X.B., Vu, D.G., Le, D.N., 2005. Ensiling of mulberry foliage (Morus alba) and the nutritive value of mulberry foliage silage for goats in central Vietnam. Livest. Res. Rural Dev. 17, 23–25.
Omar, S.S., Shayo, C.M., Uden, P., 1999. Voluntary intake and digestibility of mulberry (Morus alba) diets by growing goats. Trop. Grassl. 33, 177–181. Ørskov, E.R., McDonald, I., 1979. The estimation of protein degradability in the rumen from incubation measurements weighted according to rate of passage. J. Agric. Sci. (Cambridge) 92, 499–503. Sanchez, M.D., 2000. Mulberry: an exceptional forage available almost worldwide. World Anim. Rev. 93, 1–21. SAS, 1999. Statistical Analytical System (SAS) Users Guide: Statistics, Version 8.2.. Statistical Analysis Institute, Cary, NC, USA. Silanikove, N., Gilboa, N., Nitsan, Z., 1997. Interactions among tannins, supplementation and polyethylene glycol in goats given oak leaves: effects on digestion and food intake. Anim. Sci. 64, 479–483. Singh, B., Makkar, H.P.S., 2000. The potential of mulberry tree foliage as an animal feed supplement in India. In: Proceedings of the Electronic Conference. Food and Agriculture Organization of the United Nations, /http://www.fao.org/ag/AGA/AGAP/FRG/MULBERRY/Papers/papers. htmS. Tamminga, S., Van Vuuren, A.M., Van der Koelen, C.J., Ketelaar, R.S., Van der Togt, P.L., 1990. Ruminal behavior of structural carbohydrates, non-structural carbohydrates and crude protein from concentrate ingredients in dairy cows. Neth. J. Agric. Sci. 38, 513–526. Woods, V.B., Moloney, A.P., O’Mara, F.P., 2003. The nutritive value of concentrate feedstuffs for ruminant animals. Part II: In situ ruminal degradability of crude protein. Anim. Feed Sci. Technol. 110, 131–143.
Contents lists available at SciVerse ScienceDirect
Livestock Science journal homepage: www.elsevier.com/locate/livsci
Short communication
Nutrient composition, polyphenolic contents, and in situ protein degradation kinetics of leaves from three mulberry species W.X. Wang, H.J. Yang n, Y.K. Bo, S. Ding, B.H. Cao State Key Laboratory of Animal Nutrition, College of Animal Science and Technology, China Agricultural University (CAU), Beijing 100193, PR China
a r t i c l e i n f o
abstract
Article history: Received 13 February 2012 Received in revised form 12 March 2012 Accepted 19 March 2012
Mulberry leaves of 14 cultivars of Morus atropurpurea Roxb (MAR), 7 cultivars of Morus alba Linn (MAL) and 8 cultivars of Morus multicaulis Perr (MMP) were assessed for their differences in chemical composition and in situ ruminal degradation. The contents of acid detergent fibre (P¼ 0.073) and ether extract (Po0.0001) ranked as: MMP4MAL4MAR. Crude protein (CP), neutral detergent fibre and polyphenolic contents of the leaves did not differ between three species. Nylon bags containing the leaves were incubated for 2, 6, 12, 24, 36, 48 and 72 h in three rumen cannulated dairy cows. Soluble CP fraction and fractional degradation rate did not differ between three species, but the potential degradable CP fraction in the leaves was greater for MAR than the others (P¼ 0.005). Effective CP degradability was consistently ranked: MAR4MAL4MMP (P¼ 0.051). Nontannin phenols and total tannins were negatively correlated with the soluble CP fraction (r¼ 0.48, Po0.01) and the potential degradable CP fraction (r¼ 0.36, Po0.05), respectively. In summary, the mulberry leaves with high protein, low fibre and tannin contents could be exceptional forage for use in ruminant animals. & 2012 Elsevier B.V. All rights reserved.
Keywords: Mulberry leaves Polyphenolic content Rumen degradation
1. Introduction Mulberry trees grow under varied climatic conditions, ranging from temperate to tropical in the world. Mulberry leaves are highly palatable and rich in protein (Kandylis et al., 2008), but they also have some anti-nutritional factors, such as tannins (Makkar, 2003). Chemical composition and biomass yield of mulberry leaves vary due to the variety, maturity degree, leaf position within the branch, and fertilisation level (Sanchez, 2000). There are at least 15 species of mulberry known in China, but only the leaves of a few species have been analysed for their phytochemical profiles instead of their nutritive values (Han, 2006). In the present study, mulberry leaves from
Abbreviations: ADF, Acid detergent fibre; CP, Crude protein; DM, Dry matter; NDF, Neutral detergent fibre n Corresponding author. Tel.: þ86 10 6273 3124; fax: þ 86 10 6273 4859. E-mail address: [email protected] (H.J. Yang). 1871-1413/$ - see front matter & 2012 Elsevier B.V. All rights reserved. http://dx.doi.org/10.1016/j.livsci.2012.03.009
29 cultivars from three species, grown under the same climatic and geographic conditions, were harvested in Guangdong province. Since no studies have yet examined for digestibility values of these leaves, the objective of the present study was firstly to assess the nutritive value of these leaves in terms of chemical composition and in situ ruminal protein degradation. Secondly, we attempted to explore the relationships between the polyphenolic contents and the in situ degradation.
2. Materials and methods 2.1. Sample collection Mulberry leaves from 14 cultivars of Morus atropurpurea Roxb (MAR), 7 cultivars of M. alba Linn (MAL) and 8 cultivars of M. multicaulis Perr (MMP), were gathered during the summer season of May and June at the mulberry tree experimental plantation field of South Agricultural University
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W.X. Wang et al. / Livestock Science 146 (2012) 203–206
of China (Guangzhou, Guangdong, China) with an average altitude of 15 m above sea level. Average annual temperature was 21.9 1C, with an average minimum temperature of 13 1C in January, and a maximum medium temperature of 28.5 1C in July. Average annual precipitation was 1800 mm. The fresh leaf samples (400 g) of 5 trees per cultivar were randomly picked at the field site from the canopy at 1.0 m above the ground. The leaf samples from the same cultivar of mulberry trees were pooled and mixed to create 2.0 kg leaf samples. The samples were dried at 65 1C for 48 h and then ground to pass through a 1.18-mm screen for chemical analysis, and a 2 mm screen for in situ rumen incubation. 2.2. Animals and diet Three lactating multiparous Holstein cows, fitted with ruminal cannulae (Type 2c; Bar Diamond Inc., Parma, Idaho, USA), served as experimental animals for in situ rumen incubation. The cows had free access to water and were fed 25 kg total mixed ration divided into two equal meals at 07:00 and 17:00. The ration contained (per kg of dry matter (DM)): 250 g maize silage, 167 g ryegrass hay, 83 g alfalfa hay, 267 g maize meal, 69 g wheat bran, 138 g soybean meal, 11 g limestone, 5.5 g dicalcium phosphate, 4.5 g sodium chloride and 5 g commercial vitamin and trace mineral premix. Net energy for lactation and crude protein (CP) content was 6.69 MJ and 160 g, respectively, per kg DM. 2.3. Rumen incubations Throughout the experimental period, nylon bags (12 8 cm2 i.d.; 48 mm of pore size) were filled with 3 g leaf samples. The bags containing the leaves were sealed using rubber bands, bound to plastic strings and soaked in tap water for 20 min prior to ruminal incubation. Each cultivar leaf was incubated in each cow for the testing incubation time period: 2, 6, 12, 24, 36, 48 and 72 h after the morning feeding. After removal from the incubation, the bags were washed with tap water for 15 min until the washing ran clear and colourless. The bags were dried at 65 1C for 48 h, weighed and then the residues in the bags were used for analysis of CP content. Crude protein disappearance was calculated as the difference between the original incubated CP and the residual CP. Rumen degradation kinetics for CP disappearance per mulberry leaf per cow was calculated using the nonlinear model proposed by Ørskov and McDonald (1979). 2.4. Chemical analysis Representative leaf samples were analysed for content of DM (method 930.5), CP (method 984.13), ether extract (method 920.30), neutral detergent fibre (NDF, method 2002.04, without heat stable amylase), acid detergent fibre (ADF, method 973.18) and ash (ID 942.05) with AOAC methods (2000). Both NDF and ADF were corrected for residual ash content. Leaf samples (200 mg) were weighed into 10 ml 0.70 (v/v) aqueous acetone solution, and then were extracted with ultrasonicator at 4 1C, centrifuged at 3000 g for 20 min. Total phenolic contents in the supernatants were determined
at 725 nm by Folin-Ciocalteau reagent using tannic acid (Sigma Aldrich, St. Louis, USA) as a standard. Total tannin content was measured as the difference between total phenols before and after tannin removal by adsorption on insoluable polyvinylpyrrolidone. Non-tannin phenols content in the leaves was calculated by subtracting total tannins from total phenols content. 2.5. Statistical analyses Chemical composition measurements per mulberry cultivar were averaged before the statistical analysis. A total of 29 mulberry cultivar averages, grouped into MAR (n¼ 14), MAL (n¼7), and MMP (n¼8), were used, and statistical analyses were performed using the general linear model (SAS, 1999): Yij ¼ m þMi þ ej where Y is the dependent variable, m is the overall mean, Mi is the fixed effect of mulberry specie (i¼MAR, MAL, MMP), and ej is the random residual error. In situ degradation of the leaves with three animal replicates resulted in a total of 87 observations. After the observations per mulberry cultivar was averaged within three animals, a total of 29 mulberry cultivar averages, grouped into MAR (n¼14), MAL (n¼7), and MMP (n¼8), were used for the statistical analysis with the same model as above. The means were compared using a multiple comparison test (Tukey). The relationships between polyphenolic contents and in situ degradation kinetics were examined by a correlation analysis (SAS, 1999). Significance was declared at Po0.05 unless significance P values between 0.05 and 0.10 are considered as trends and reported. 3. Results and discussion Mulberry leaf are of a high protein quality comparable to or better than soybean meal (Machii, 1989), and it could be an alternative supplemental feed, comparable and in some cases superior to alfalfa, to replace conventional feed resources for goat (Nguyen et al., 2005) and sheep (Doran et al., 2007). In the present study, CP and NDF contents did not differ between three species (Table 1). Ether extract (Po0.0001) and ADF (P¼0.073) contents ranked as: MMP4 MAL4MAR. The CP content was close to the upper value noted in the literature (180–250 g/kg DM; Fotadar and Dandin, 1997; Omar et al., 1999). Feeds with similar CP availability in the rumen could be used to synchronise nutrient supply for maximum microbial protein yield (Herrera-Saldana et al., 1990). In the present study, the highest potential degradable CP fraction occurred in MAR (P¼0.005), and no differences for soluble CP fraction and fractional degradation rate were observed in the leaves between three species (Table 1). Effective CP degradability ranked as: MAR4MAL4MMP (Po0.051). Compared to the results reported by Woods et al. (2003), the soluble CP fraction in the present study was lower than most protein feeds (e.g., soybean meal and maize gluten); the potential degradable CP fraction was higher; the rate was similar or somewhat higher, and the effective degradability was similar when the passage rate k was assumed as 0.08/h. The degradation rate and effective CP degradability in the present study were quite close to those of MAL leaves observed by Singh and Makkar (2000), who noted that the
W.X. Wang et al. / Livestock Science 146 (2012) 203–206
205
Table 1 Chemical composition and ruminal crude protein degradation kinetics of leaves from Morus atropurpurea Roxb (MAR), Morus alba Linn (MAL) and Morus multicaulis Perr (MMP). MAR
MAL
MMP
S.E.M
P value
Chemical composition (g/kg dry matter) Crude protein Ether extract Neutral detergent fibre Acid detergent fibre Total phenolics Total tannins Non-tannin phenols
251.7a 28.5c 278.8a 164.9b 15.5a 1.2a 14.3a
259.0a 42.1b 262.5a 170.7ab 14.3a 1.3a 13.0a
251.8a 49.1a 275.4a 176.6a 15.2a 1.6a 13.6a
4.03 1.84 5.89 3.00 0.58 0.20 0.44
0.531 o0.0001 0.279 0.073 0.502 0.495 0.239
Rumen degradation kinetics Soluble protein Potential degradable protein Fractional degradation rate Effective degradability
0.06a 0.94a 0.048a 0.43a
0.09a 0.90b 0.046a 0.40b
0.07a 0.89b 0.043a 0.36c
0.014 0.010 0.0027 0.012
0.323 0.005 0.341 0.051
In the table, rumen degradation kinetics were calculated using the nonlinear model proposed by Ørskov and McDonald (1979): y¼aþb (1 e c t) where y¼ degradability for crude protein at time t relative to incubation (h), a¼ soluble fraction, b¼ potential degradable fraction, and c¼fractional degradation rate. Effective degradability was calculated using the equation: ED¼ aþ (bþc)/(cþk) with an assumed passage rate (k) of 0.08/h (Tamminga et al., 1990). Means in a row without a common superscript differ as noted P value.
kinetic parameters of CP in their mulberry leaves were comparable to those of other good quality tree fodders such as Leucaena leucocephala and Ficus roxburghii. As reviewed by McSweeney et al. (2001), tannins are polyphenolic compounds of plant origin, which are of two distinct types, hydrolysable tannins (polyesters of gallic acid and various individual sugars) and condensed tannin (polymers of flavonoids). Condensed tannin content ( o27 mg/kg DM) measured in the present study may be negligible for the mulberry leaves and was not presented in Table 1. No distinct differences occurred for total phenols, total tannins, non-tannin phenols in the leaves between three species (Table 1) in which the total phenols contents in the leaves were slightly less than 18 g/kg DM reported by Singh and Makkar (2000). The presence of tannins in nutritionally important forage trees, shrubs, legumes, cereals and grain often limits their feed utilisation, as evidenced by Silanikove et al. (1997). The tannin compounds can have toxic and/or anti-nutritional effects on animals due to the complexing (tanning) ability with dietary protein, which then inhibits the growth of microorganisms (McSweeney et al., 2001). Non-tannin phenols and total tannins in Table 2 were negatively correlated with the soluble CP fraction (r¼ 0.48, Po0.01) and the potential degradable CP fraction (r¼ 0.36, Po0.05), respectively. These findings were consistent with Liener’s study (1980), in which tannins caused an adverse effect on protein digestibility.
Table 2 Partial correlation coefficients between polyphenolic contents in mulberry leaves (n¼ 29 cultivars) and in situ crude protein degradation kinetics. Total phenol
Total tannins
Non-tannin phenols
Soluble protein 0.42n Potential degradable protein 0.19 Fractional degradation rate 0.36n
0.12 0.36n 0.37n
0.48nn 0.02 0.24
P o0.01. P o 0.05.
nn n
Conflict of interest statement We respectfully submit our manuscript entitled ‘‘Nutrient composition, polyphenolic contents, and in situ protein degradation kinetics of leaves from three mulberry species’’ for your consideration for publication in the Livestock Science, Section of Ruminant Nutrition. All authors agree to this submission and affirm that the manuscript presents original work by the authors and is not being considered by any other journal. Hereby, we want to state that there are no any conflict of interests for the submission of this manuscript.
Appendix A. Supplementary material 4. Conclusion The mulberry leaves appear to have a high potential as a protein-rich forage with low fibre content and could serve as comparatively less degradable protein in the rumen. Although there may be associations between polyphenolic compound deposition and degradability measures that cannot be accounted for through simple correlation analysis, mulberry leaves with very low tannin content could be exceptional forage for ruminant animals.
Supplementary data associated with this article can be found in the online version at doi:10.1016/j.livsci.2012. 03.009.
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