Quality of alfalfa (Medicago sativa L.) and red clover (Trifolium pratense L.) mixture silages depending on the share in the mixture and additives


  • Đorđe LAZAREVIĆ Institute for Forage Crops, 37251 Globoder, Kruševac (RS)
  • Vladeta STEVOVIĆ University of Kragujevac, Faculty of Agronomy, Cara Dušana 34, 32000 Čačak (PT)
  • Zoran LUGIĆ Institute for Forage Crops, 37251 Globoder, Kruševac (RS)
  • Dalibor TOMIĆ University of Kragujevac, Faculty of Agronomy, Cara Dušana 34, 32000 Čačak (RS)
  • Jordan MARKOVIĆ Institute for Forage Crops, 37251 Globoder, Kruševac (RS)
  • Vladimir ZORNIĆ Institute for Forage Crops, 37251 Globoder, Kruševac (RS)
  • Mladen PRIJOVIĆ Institute for Forage Crops, 37251 Globoder, Kruševac (RS)




alfalfa , inoculation, mixture, red clover, silage, tannin


Alfalfa silage has a high potential for improving ruminant nutrition. A problem that frequently occurs during its preparation is the process of proteolysis, which could partially be avoided with the use of certain techniques and materials. Red clover, often used in form of silage, expresses weaker proteolysis due to the possession of certain chemical compounds. This research was conducted to study the effects of ensiled alfalfa and red clover mixtures, mixed at different ratios (100:0, 90:10, 70:30, 50:50, 30:70, and 0:100), as well as the influence of additives – two doses of oak tannin extract (6 g kg-1 DM and 12 g kg-1 DM) and bacterial inoculant (Enterococcus faecium, Bacillus plantarum, and Bacillus brevis) on fermentation parameters and protein fractions of the silages. The treatments which contained any of the used percentages of red clover, as well as the ones which received the higher dose of oak tannin (12 g kg-1 DM), reduced the ammonia nitrogen content, which is one of the main indicators of proteolysis. On the other hand, considering non-protein nitrogen, as another of the indicators of proteolysis, there was no positive sign of the contribution of red clover to the reduction of proteolysis in alfalfa, and neither there was a significant impact of additives that were applied in this experiment.


Albrecht KA, Muck RE (1991). Proteolysis of ensiled forage legumes that vary in tannin concentration. Crop Science 31:464-469. https://doi.org/10.2135/cropsci1991.0011183x003100020048x

AOAC (1990). Official method 984.13 Crude protein in animal feed, forage, grain, and oil seeds. Official methods of analysis of AOAC International, 15th edition.

Broderick GA (1995). Performance of lactating dairy cows fed either alfalfa silage of alfalfa hay as the sole forage. Journal of Dairy Science 78:320-329. https://doi.org/10.3168/jds.S0022-0302(95)76640-1

Dinić B, Đorđević N, Lugić Z, Lazarević D, Marković J (2006). Lucerne and red clover conservation by method of ensiling. 17th symposium on innovation in animal science and production. Biotechnology in Animal Husbandry 22:535-548.

Dong Z, Chen L, Li J, Yuan X, Shao T (2019). Characterization of nitrogen transformation dynamics in alfalfa and red clover and their mixture silages. Grassland Science 65:109-115. https://doi.org/10.1111/grs.12230

Dong Z, Yuan X, Wen A, Desta ST, Shao T (2017). Effects of calcium propionate on the fermentation quality and aerobic stability of alfalfa silage. Asian-Australasian Journal of Animal Sciences 30:278-284. https://doi.org/10.5713/ajas.16.0956

Đorđević N, Dinić B (2003). Siliranje leguminoza [Ensiling of legumes]. Monografija. Institut za istraživanja u poljoprivredi, Srbija.

Fijałkowska M, Pysera B, Lipiński K, Strusińska D (2015). Changes of nitrogen compounds during ensiling of high protein herbages: a review. Annals of Animal Science 15(2):289-305. https://doi.org/10.1515/aoas-2015-0008

Fox DG, Tedeschi LO, Tylutki TP, Russell JB, Van Amburgh ME, Chase LE ... Overton TR (2004). The Cornell Net Carbohydrate and Protein System Model for evaluating herd nutrition and nutrient excretion. Animal Feed Science and Technology 112(1-4):29-78. https://doi.org/10.1016/j.anifeedsci.2003.10.006

Guo XS, Ding WR, Han JG, Zhou H (2008). Characerization of protein fractions and amino acids in ensiled alfalfa treated with different chemical additives. Animal Feed Science and Technology 142:89-98. https://doi.org/10.1016/j.anifeedsci.2007.07.005

Herremans S, Decruyenaere V, Beckers Y, Froidmont E (2019). Silage additives to reduce protein degradation during ensiling and evaluation of in vitro ruminal nitrogen degradability. Grass and Forage Science 74:86-96. https://doi.org/10.1111/gfs.12396

Higgs RJ, Chase LE, Ross DA, Van Amburgh ME (2015). Updating the cornell net carbohydrate and protein system feed library and analyzing model sensitivity to feed inputs. Journal of Dairy Science 98:6340-6360. https://doi.org/10.3168/jds.2015-9379

Huo W, Wang X, Wei Z, Zhang H, Liu Q, Zhang S ... Guo G (2021). Effect of lactic acid bacteria on the ensiling characteristics and in vitro ruminal fermentation parameters of alfalfa silage. Italian Journal of Animal Science 20(1):623-631. https://doi.org/10.1080/1828051x.2021.1906167

Jones BA, Hatfield RD, Muck RE (1995). Characterization of proteolysis in alfalfa and red clover. Crop Science 35:537-541. https://doi.org/10.2135/cropsci1995.0011183x003500020043x

Krawutschke M, Weiher N, Gierus M, Thaysen J (2011). The effect of cultivar on the crude protein fractions of fresh, wilted and ensiled red clover. Grassland Science in Europe 16:256-258.

Li X, Tian J, Zhang Q, Jiang Y, Wu Z, Yu Z (2018). Effects of mixing red clover with alfalfa at different ratios on dynamics of proteolysis and protease activities during ensiling. Journal of Dairy Science 101:1-11. https://doi.org/10.3168/jds.2018-14763

Licitra G, Hernandez TM, Van Soest PJ (1996). Standardization of procedures for nitrogen fractionation of ruminants feeds. Animal Feed Science and Technology 57:347-358. https://doi.org/10.1016/0377-8401(95)00837-3

Marković J, Milenković J, Anđelković S (2021). Alfalfa and red clover as a protein source for ruminants. Proceedings of the 13th International Symposium Modern Trends in Livestock Production October 6-8, 2021, Belgrade, Serbia pp 308-321.

Mayer AM (1986). Polyphenol oxidases in plants – Recent progress. Phytochemistry 26:11-20. https://doi.org/10.1016/S0031-9422(00)81472-7

McDonald P, Henderson AR, Heron SJE (1991). The Biochemistry of Silage. 2nd Edition, Chalcombe Publications, Marlow, UK pp 340. https://doi.org/10.1017/S0014479700023115

Muck RE, Bolsen KK (1991). Silage preservation and silage additives. In: Bolsen KK, Baylor JE, McCullough ME (Eds). Hay and Silage Management in North America. National Feed Ingredients Association. West Des Moines. Iowa pp 105-125.

Njegić R, Žižić M, Lovrić M, Pavličić D (1991). Osnovi statističke analize [Basics of the statistical analysis]. III izdanje, Savremena administracija, Beograd pp 498.

Ohshima M, McDonald P (1978). A review of the changes in nitrogenous compounds of herbage during ensilage. Journal of the Science of Food and Agriculture 29:497-505. https://doi.org/10.1002/jsfa.2740290602

Owens VN, Albrecht RE, Muck RE, Duke SH (1999). Protein degradation and fermentation characteristics of red clover and alfalfa silage. Crop Science 39:1873-1880. https://doi.org/10.2135/cropsci1999.3961873x

Purwin C, Fijałkowska M, Kowalik B, Nogalski Z, Pysera B (2014). The effect of bale density and addition of formic acid on the in situ dry matter and crude protein degradation of lucerne, red clover and red fescue silages. Journal of Animal and Feed Sciences 23(2):177-184. https://doi.org/10.22358/jafs/65707/2014

Sniffen CJ, O'Connor J, Van Soest P, Fox D, Russell J (1992). A Net Carbohydrate and Protein System for evaluating cattle diets: II. Carbohydrate and protein availability. Journal of Animal Science 70:3562-3577. https://doi.org/10.2527/1992.70113562x

StatSoft Inc. (2007). STATISTICA (data analysis software system), version 8.0.www.statsoft.com.

Sullivan ML, Hatfield RD (2006). Polyphenol oxidase and o-diphenols inhibit postharvest proteolysis in red clover and alfalfa. Crop Science 46:662-670. https://doi.org/10.2135/cropsci2005.06-0132

Tao L, Zhou H, Zhang N, Si B, Tu Y, Ma T, Diao Q (2016). Effects of different source additives and wilt conditions on the pH value, aerobic stability, and carbohydrate and protein fractions of alfalfa silage. Animal Science Journal 88:99-106. https://doi.org/10.1111/asj.12599

Van Amburgh ME, Recktenwald EB, Ross DA, Overton TR, Chase LE (2007). Achieving better nitrogen efficiency in lactating dairy cattle: Updating field usable tools to improve nitrogen efficiency. In: Proceedings of 18th Cornell Nutrition Conference, 30-31st January, Syracuse, NY pp 25-38.

Wiegner G (1926). Anleitung zum qaantitativenagrikulturchemischen Praktikum [Instructions for the quantitative agricultural chemical practical course]. Berlin, Gebriider, Borntraeger.



How to Cite

LAZAREVIĆ, Đorđe, STEVOVIĆ, V., LUGIĆ, Z., TOMIĆ, D., MARKOVIĆ, J., ZORNIĆ, V., & PRIJOVIĆ, M. (2023). Quality of alfalfa (Medicago sativa L.) and red clover (Trifolium pratense L.) mixture silages depending on the share in the mixture and additives. Notulae Botanicae Horti Agrobotanici Cluj-Napoca, 51(1), 12954. https://doi.org/10.15835/nbha51112954



Research Articles
DOI: 10.15835/nbha51112954

Most read articles by the same author(s)