Only a few previous studies have analysed the crude protein (CP) fractions of the Cornell Net Carbohydrate and Protein System during the growth period of forage legumes. The objective of the present study was to investigate the changes in CP fractions during the spring growth and summer-autumn regrowth period of five forage legume species (alfalfa (also known as lucerne, Medicago sativa L.), birdsfoot trefoil (Lotus corniculatus L.), kura clover (Trifolium ambiguum M.B.), red clover (Trifolium pratense L.) and white clover (Trifolium repens L.)) grown in binary mixtures with perennial ryegrass (Lolium perenne L.) and also in pure stands (two red clover cultivars). Additionally, the specific polyphenol oxidase (PPO) activity was measured photometrically in the leaves of pure red clover swards. In both pure and mixed cropping, CP fraction A increased with advancing maturity, except for the legumes from mixed cropping in the summer-autumn growth period 2004 and 2005. The variation of CP fraction A was mostly positively related to the N yield and the amount of dinitrogen fixation. Although CP fraction A of pure red clover was negatively correlated with the specific PPO activity in the spring growth period, the specific PPO activity was less relevant for the variation of CP fraction A with respect to the whole growing season. CP fraction B generally made up the largest proportion of the CP. Pure red clover stands showed reducing amounts of CP fraction C during the growth period, whereas in legumes grown with ryegrass an increase was usually observed. Despite these differences, there was generally an increase of CP fraction C when the content of nonstructural carbohydrates decreased. Red clover and birdsfoot trefoil herbage contained the highest proportions of CP fraction C in the CP, regardless of growth period and year. In conclusion, red clover and birdsfoot trefoil had a more favourable CP composition for ruminant nutrition compared to the other legume species, and in red clover this could not be clearly attributed to the specific PPO activity.

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Various management practices (e.g. wilting, application of silage additives or adding a grass component) can be used to improve silage fermentation of pure red clover (Trifolium pratense L.). Therefore, the aim of this laboratory ensiling study was to investigate the effects of varying proportions of red clover and perennial ryegrass (100/0, 66/33, 33/66, 0/100) on silage quality during two consecutive years. In addition, two wilting levels [target dry matter (DM): 300 vs. 400 g kg−1] in combination with lactic acid bacteria (LAB) additives were tested. Herbage was ensiled, either untreated or inoculated with homofermentative LAB (low wilted) or homo‐ and heterofermentative LAB (high wilted). In most cases, lactic and acetic acid decreased as the proportions of ryegrass were increased. Data concerning ammonia‐N concentrations showed considerable differences between cuts and years. Silages treated with homofermentative LAB generally had high lactic acid and low final pH, whereas acetic acid and 1,2‐propanediol tended to be higher when homo‐ and heterofermentative LAB were applied. Inoculants had a positive effect on DM losses and ammonia‐N in only a few silages. Wilting decreased DM losses and fermentation acids at most cuts, irrespective of the grass/clover ratio in the herbage mixture. There was a strong year effect on the organic matter digestibility (DOM) of the silages. In conclusion, the optimal strategy for successful silage fermentation of red clover is the ensiling in mixtures with ryegrass. Furthermore, herbage should be wilted to a DM content of about 300–350 g kg−1. The application of LAB inoculants did not alter the DOM but did improve silage fermentation.

SUMMARYRed clover (Trifolium pratense L.) silage usually contains lower contents of non-protein nitrogen (NPN) compared with other forage legumes. This is often attributed to the polyphenol oxidase (PPO) activity in red clover, although in most field studies the PPO activity was not measured. Therefore, a laboratory ensiling experiment with three red clover cultivars and one white clover cultivar as control grown in two management systems (with and without mechanical stress) over 2 consecutive years was conducted. Fresh, wilted and ensiled clover herbage was sampled at four cutting dates per year to determine the crude protein (CP) fractions according to the Cornell Net Carbohydrate and Protein System. The specific PPO activity was measured photometrically in fresh clover leaves. The content of CP fraction A (NPN) increased from fresh over wilted to ensiled clover herbage at the expense of the content of CP fraction B (true protein), irrespective of species, cultivar and year. The most important source of variation for all CP fractions and the calculated rumen-undegradable protein contents was generally the herbage condition, except for CP fraction C (unavailable protein). White clover silage consisted of higher contents of CP fraction A and lower contents of CP fraction B3 in CP compared with red clover silage. As a result, the calculated rumen-undegradable protein content of white clover silage was lower than that of all red clover cultivars. In conclusion, the extent of proteolysis during ensiling among the silages made from the herbage of different red clover cultivars was primarily influenced by the stage of maturity at harvesting and the degree of wilting at ensiling. The variation in specific PPO activity could not be related to the extent of proteolysis.