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Nutritive value, and in situ dry matter and protein degradability of fresh and ensiled Italian ryegrass at three stage of growth


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BORBA, ALFREDO E.S. & JoAO M.C. RAMALHO RIBEIRO 1996. Nutritive value,

and in situ dry matter and protein degradability of fresh and ensiled Italian

ryegrass at three stage of growth. Arquipe'lago. Life and Marine Sciences 14A: 85-

94. Ponta Delgada. ISSN 0873-4704.

The nutritive value of Italian ryegrass (Lolium multiflorum) cut at three stages of growth and in fresh and ensiled, was determined in this study. The highest metabolisable energy content was found at 5% ear in fresh forages, and at the fourth leaf stage in silages. In vivo digestibility and vonluntary intake increased from fourth leaf to the 5% ear stage of growth which was followed by a reduction at the 100% ear stage of growth. In situ dry matter and protein degradability decreased (p10.05) with advancing stage of growth. In fresh forages protein degradability was higher (p10.05) than in ensiled forages. It was concluded that the best harvest time for Italian ryegrass was at the 5% ear stage of growth for both fresh forage, as well as silage.

BORBA, ALFREDO E.S. & JOAO M.C. RAMALHO RBEIRO 1996. Variaqgo do valor

nutritivo e da degradabilidade


situ da mattria seca e da proteina bruta da erva

castelhana corn a 6poca de corte e corn o mttodo de conserva@o. Arquipe'lago.

CiEncias Biol6gicas e Marinhas 14A: 85-94. Ponta Delgada. ISSN 0873-4704.

Neste trabalho determiniimos o valor nutritivo da erva castelhana (Lolium multflorum) em

t r b tpocas de corte, em verde e ensilada. 0 valor mais elevado de energia metabolizdvel foi determinada na fase de crescimento 5% espigado, nas forragens verdes e na fase de aparecimento da 4" folha, para as silagens. Verificou-se um aumento da digestibilidade in vivo e da ingest20 volunt&ria, da fase de 4" folha para a fase 5% espigada, seguindo-se urn decrtscimo para a fase 100% espigada: A degradabilidade in siru da matiria seca, e da

proteina bruta decresce (pS0.05) corn o avancar da fase de crescimento. A degradabilidade

da proteina bruta 6 superior nas forragens verdes (p10.05) em relac20 ios forragens ensiladas. Podemos concluir que a melhor 6poca de corte, para a erva castelhana, verifica- se na fase de crescimento com 5% espigado.

Alfredo E.S. Borba, Universidade dos A~ores, Departamento de Ci2ncias Agrdrias, PT- 9700, Angra do Heroismo, Agores, Portugal.


Jorlo M. C. Ramallto Ribeiro, Esta~rlo ZootPcnica Nacional, Departamento de Nutrig80, PT-2000 Vale de SantarPm, Portugal.

INTRODUCTION ensiled under the climatic conditions in the

Azores. TISSERAND (1984) reported that

5% ear

Italian ryegrass (Lolium multi$!orum) is one of the stage of growth is the highest production of

more temporary pastures utilised in the Azores. digestible dry matter.

The present research was carried out to determine The

in siru

technique has been used for many

the nutritive value of this forage at three stages of years to determine the dry matter digestibility of


This method represents an approach to the and fatty acids by gas chromatography using the digestive process. MEHREZ & QRSKOV (1977) method of JOUANY (1981).

have also used this procedure to measure protein degradability.

IN VIVO DIGESTIBILITY The present experiment was undertaken to

determine the dynamically dry matter

digestibility; the digestibility of fresh and ensiled Italian ryegrass. A second objective was to determine the ill situ protein degradability of the Italian ryegrass forage.


The Italian ryegrass (Loliurn mulriflorum, L.), was the Multimum variety, and was harvested at three stages of growth: appearance of the fourth leaf (L); with 5% ear (E); and 100% ear (H), and it was analysed in the fresh (F) and ensiled (E) form.

Forages were harvested with a precision-chop machine with a theoretical chopping length of 15 mm. Approximately 1.5 tones of each fresh forage was frozen at a temperature of -15"C, and other portion (also 1.5 tones approximately) was ensiled. The silos were opened after 60 days.

Forage was dried at 65"C, and ground through a 1.0 mm screen. The following parameters were measured: dry matter (DM); ash; crude protein (CP); ether extract (EE) and crude fiber (CF) for the Weende system (AOAC 1975). Gross energy was determined in adiabatic bomb. The forages were also subject to the following analyses: NDF. ADF and ADL (GOERING & VAN SOEST 1970), MADF (CLANCY & WILSON 1966), and calcium and phosphorous (OJEC 1971).

The following were quantified in fresh silages: pH; N-NH3 by the Conway method (CONWAY 1957); water soluble carbohydrates by the Deriaz method (DERIAZ 1961) and ethanol. lactic acid

Six adult Romney-Marsh male sheep, per treatment, were used for the in vivo digestibility determinations.

Digestibility trials lasted 21 days; there was a 14 day adaptation period and a 7 day collection period. Animals were fed ad libitunz twice daily. at 09.00 a.m. and 17.00 p.m. Quantities of offered feed were the previous days consumption amounts plus 10%.

Urines were acidified with concentrated hydrochloric acid (HCI) during collection, in order to prevent the losses of the volatile compounds.

Digestible energy (DE) and metabolisable energy (ME) has been determined from the values of crude energy (CE) ingested and faecal and urinary excretions obtained with i r ~ vivo assays of digestibility. To determine the metabolisable energy, we selected a value of 7% for the loss of energy as methane gas form (MCDONALD et al.


Three adult Romney-Marsh male sheep, fitted with permanent 40 mm cannula were used to measure the in situ degradability. They were fed a standard mean quality hay diet ad libitunz along with 200 concentrate and mineral supplement.

Nylon bags made of precision woven nylon cloth (Hydro-Bios- NY55 HC) measuring 17 x 9 cm and with a pore size of 55 pm were used for the in situ method. Approximately 5 g of the sample were placed in each bag. Three bags were prepared for each treatment. Bags were attached to a PVC structure. These were attached to the fistula cap with a 25 cm long nylon string. Bags were immersed completely in water to acquire weight and were incubated in the rumen. W e


have incubated 6 bags in each sheep. Bags were incubated for 0, 1, 3, 6, 12, 24, 48 and 72 hours. The zero-hour bags were only washed in running water for 15 minutes. When taken out of the rumen, bags were washed under running water, until rinsing water was colourless and dried at 65°C in a ventilated oven. The weight and crude protein content of the residues were then determined.

We determined the fractional outflow (k) of the particleslhour from the rumen, by treating 50 g of basal diet with sodium-dichromate technique (UDEN et al. 1979). Following the method described by GANEV et al. (1979), the basal diet was added through a cannula to the rumen of two sheep, which received a standard hay diet ad libitlim. Samples of fresh rumen contents were obtained through the cannula at I , 2. 4, 6, 9, 12, 15, 18. 21, 24, 36 and 48 hours after the addition of the material. Samples were dried at 105°C for determination of dry matter. The method described by STEVENSON & LANGEN (1960) was used for chromium determination. The amount of chromium in the rumen liquor were fitted to the equation:

X= e-kt (1)

in which X represents the chromium fraction that remains in the rumen after the time t and k is the fractional outflow rate (HUNGATE 1966).

Dry matter and crude protein degradability (P) at each one of a series of incubation times (t) is exponential and can be described, following ~ R S K O V & MCDONALD (1979), by the equation:

in which a is the measure of the rapidly soluble dry matter or protein fraction, b the fraction which is subjected to degradation and c the constant rate of disappearance of fraction b. The constant a, the constants c and the potential dry matter and protein degradability (a+b) were determined by the method of MCDONALD (1 98 1).

The effective dry matter degradability (P) was calculated, following ~ R S K O V & MCDONALD (1979) from the relationship:


The experimental data were analysed as a 3x2 factorial with six repetitions, with three stages of growth, for fresh and ensiled materials using multiple comparison of the average means were computed using the t-test (STEEL & TORRIE


RESULTS AND DISCUSSION The chcmical composition of the forage is give in tables 1 and

Italian ryegrass 2. Dry matter


content increased with advancing of the stage of growth, with a high reduction of crude protein content. The crude protein content of the Italian ryegrass (134 to 27 1 glkg) was much higher than found in literature.

The fiber, or the cell wall fraction, represented by crude fiber, NDF, ADF, MADF and ADL, increased with advancing of stage of growth.

Based on thc pH values and the classification used by VANBELLE et al. (1981), the LS and ES silage can be classified as average quality silage whereas the HS was a poor silage. Based in N- NH3 and followed the same authors, the LS silage can be considered acceptable, the ES good, the HS a poor silage.

Based on other observations, the LS silage could be classified as an acetate silage, the ES as a lactate silage and the HS as an acetate silage, with higher content in butyric acid.

The results obtained were a surprising; the HS forage had a high content of water soluble carbohydrates when compared with other forages and ones reported by MCGRATH (1992). MULLER (1981) obtained good results with an Italian ryegrass silage in an advanced stage of maturity (i.e. 27% of DM). The lower quality of the LS silage can be explained by its lower content in water soluble carbohydrates of the fresh forage.


Table 1

Mean chemical composition of the green and ensiled Italian ryegrass (n=3): all values are presented in gkg DM except DM which represents glkg material.





Ash WSC Ca P


119.0 256.3 204.1 498.6 264.1 252.7 9.5 15.5 168.2 9.5 49.6 3.3

HS 142.0 134.4 278.4 636.3 406.5 387.2 28.3 17.9 103.6 0.6 38.1 2.0

DM= Dry matter; CP= Crude protein; CF= Crude fiber; ADF= Acid detergent fiber; NDF= Neutral detergent fiber;

ADL= Acid detergent lignin; MADF= Modified acid detergent fiber; WASC= water soluble carbohydrate; EE=

Ether extract.

Table 2

Additional mean chemical data for Italian ryegrass silages (n=3).

NH3/ Fatty acids- all in lOOg DM

Forages pH %TN lactic acetic propionic butyric ethanol

LS 5.00 13.31 0.85 5.74 0.29 0.47 0.78

The digestible and metabolisable energy contents of Italian ryegrass (Table 3) decreased (p10.05) with increasing plant age. An exception to the forages in the stage 100% ear was observed, in which the content in energy of the ensiled forages was higher than in fresh forages.

The in vivo digestibility of Italian ryegrass (Table 4) decreased with advance of the stage of growth. We observed a significantly decrease (pS0.05) between the nutritive value of the Tresh forages and silages at all stags, for the measured

parameters. MUNRO & WALTERS (1985) observed

that the Italian ryegrass is more digestible that other gramineae because they have a higher cell

content. Same authors have reported digestibility values closed to what we have determined


MICHALET-DUREAU 1983; O'KIELY et al. 1989).

Others, however have reported higher

digestibilities (LIPPKE & BARTON 1988;



Voluntary intake of fresh forage was highest in the 5% ear stage of growth. We observed an

increase in intake for EF forage and then a

reduction in the HF forage. The same tendency


Table 3

Mean energy value of green Italian rygrass and ensiled forages.

forages MJ/ Kg DM

CE* DE** ME**

CE= Crude energy; DE= Digestible energy; ME= Metabolizable energy; jC n=3;



Table 4

Mean values (n=6) of in vivo digestibility (gkg DM) and dry matter voluntary intake (g DMkg live ~ e i g h t O . ~ ~ ) o f green and ensiled Italian rygrass. Standard deviations are given in brackets.

forages DM OM D value CP NDF ADF DM Intake g/Kg 0.75

LF 682 c 728 d 606 c 695 d 850 d 796 d 49 ab ( 2 1 8 ) (+17) (+15) (+29) (+25) ( c 3 1 ) ( 2 1 0) LS 6 6 4 b c 7 1 4 d 6 0 4 c 7 4 0 e 8 4 2 d 8 2 1 e 43 a (2 56) ( 2 47) ( 2 42) (+ 44) (+ 32) (+ 37) (2 6) E F 7 1 2 d 714d 6 2 0 c 601 b 782 c 769 c 68 d ( 2 3 7 ) (+36) (+31) ( 2 2 9 ) (+24) ( ~ 1 7 ) ( 2 6) ES 649 b 683 c 600 c 624 bc 770c 760 c 64 cd (+20) (c17) ( 2 1 5 ) ( c 3 1 ) ( c 1 5 ) ( 2 1 3 ) (+ 8) HF 5 7 2 a 5 9 2 b 5 5 1 b 6 5 1 c 6 0 2 a 5 3 2 a 62c (+ 28) (+ 27) (+ 25)


34) ( 2 33) ( 2 35) (+1 1) HS 5 6 4 a 5 7 0 a 5 1 1 a 4 9 0 a 6 6 2 b 604b 52 b (222) ( 2 21) (+ 18) (+58) (+ 15) ( 2 31) ( 2 7) S Growth
















OM= Organic matter: S= level of significance 50.05; a,b.c,d and e- Where similar letters on the same line are indicated. there are not significant differences between means.

The lower intake of the young forage was is unlikely that the intake of fresh grass is probably due to a higher content of water. reduced by the higher internal water content or Influence of the water content on voluntary intake by the water from the rain in grass surface. has been reported by different authors DEMARQUILLY (1966) reported that dry matter (CAMPLING 1964; HOLMES & LANG 1963). intake significantly increased with the content HOLMES & LANG (1963) had the opinion that it in dry matter.


Constants of dry matter degradability are presented in Table 5. W e observed a decrease of the fraction b with the advancing stage of growth.

Dry matter degradability decreased

(p10.05) with advancing plant development, specially in ensiled forages (p10.05). The effective dry matter degradability estimated by equations. when k determined was 0.058/hour, also was reduced with advancing plant maturity (Fig. I).

111 situ dry matter degradability was higher in fresh than in ensiled forages (Fig. 2). These are similar to those observed in vivo digestibility.

In relation to crude protein degradability, the constants showed that the rapidly soluble fraction (a) was higher in fresh forages than in silages. W e also observed a decrease in the fraction which was subjected to degradation (b), with the development of the plant (Table 5).

Table 5

Constants of dry matter and protcin degradabilily from the fresh and ensiled Italian ryegrass forages.

Incubation time (hours) Forage

Fig. 1. Dry matter degradability, as a function of time. calculated by the


& MCDONALD (1979) cquation. P = a


b ( ~ - e - ~ ~ ) , for fresh and ensiled Italian ryegrass forages.

Dry matter

a b c rsd

Crude protein


C r u d e protein degradability determined by advancing growth stage. T h e effective crude

the equation 2 was significantly higher protein degradability found by the equation 3

(p10.05) in fresh than in silage forages (Fig. (Fig. 4), when k was 0.058/hour, had a similar

3), and decreased significantly (p10.05) with conduct.

Incubation rime (hours)

Fig. 2. Effective dry matter degradability, as a function of time. calculated by the ~ R ~ K O V & MCDONALD (1979) equation. P = a


(I-e -(cfk)t ), for fresh and ensiled Italian ryegrass forages.


Time (hours)

Fig. 3. Crudc protein dcgradability, as a function of time, calculated by the ~ R S K O V & MCDONALD (1979) equation. P = a


b (I-eFct). for fresh and ensilcd Italian ryegrass forages.


Time (hours)

Fig. 4. Effective crude protein degradability, as a function of time, calculated by the ~ R S K O V & MCDONALD (1979) equation, P = a



(I-e -(c+~)~), for fresh and ensiled Italian ryegrass forages.

The effective crude protein degradability, estimated by equation 3, was higher in fresh than

in ensiled forages.


et al. (1993)

reported that mature grasses were lower in

ruminally degradable CP. VIK-MO (1989) and



(1992) found a decrease of the nitrogen degradability, in silages, parallel with an increased DM content.


In this study we observed a higher digestibility and voluntary intake, for the Italian ryegrass at the


ear stage of growth and a reduction in the

nutritive value when with ensiled matter than fresh forages.

The best harvesting time is in the stage of

growth 5% ear, either to be utilised as fresh

forage, or as silage.

We can assume that the in situ dry matter

degradability of the fresh and ensiled Italian ryegrass forages decreases with the advance of the plant stage of growth and is higher in fresh than in ensiled forages.

In situ crude protein degradability of Italian

ryegrass is higher in ensiled than in fresh forages,

and decreasing with the advance of the stage of growth.


The authors express their appreciation to Goretti Fagundes and Gilberta Goulart for their excellent technical assistance in the laboratory.

We would like to thank Eng0 J.F. Moreira da Silva for advice and valuable discussions and for the revision of the English text.




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kco ole

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401 -406.


Fig.  2.  Effective dry matter degradability, as a function of time. calculated by the  ~ R ~ K O V   &  MCDONALD  (1979)  equation
Fig. 4. Effective crude protein degradability, as  a  function of time, calculated  by  the  ~ R S K O V   &  MCDONALD  (1979)  equation,  P =  a  +  &  (I-e -(c+~)~),  for fresh and ensiled Italian ryegrass forages


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