Effect of stage of harvest on DM yield, nutrient content, in vitro and in situ parameters and their relationship of native and Brachiaria grasses in the Adamawa Plateau of Cameroon

M B Enoh, C Kijora*, K J Peters*and S Yonkeu

Wakwa Institute of Agricultural Research for Development, P.O Box 65 Ngaoundere, Cameroon
*Institute of Animal Sciences, Humboldt-University Berlin, 10115 Berlin, Germany
claudia.kijora@rz.hu-berlin.de

Abstract

A Hyparrhenia dominated native and an introduced cultivated Brachiaria (Brachiaria ruziziensis Germain and Evrard) pasture from the Adamawa Plateau in Cameroon were evaluated for their yield and nutrient content.

The yield and quality of the introduced pasture were higher than in the native one. The quality of the pastures followed a negative trend as the length of growing period prolonged from 8 to 12 weeks. In situ dry matter degradation parameters were fitted to the exponential model of DM disappearance (Y = a + b (1 - exp ^-ct ). The potential degradability (PD) value was correlated with estimated chemical parameters, enzymatic cellulose solubility and the washing loss from nylon bags (2 hours soaking, 39°C). The best method for estimation of digestibility in this investigation seems to be the water solubility. Considering DM yield and total nutrient yield the optimum time of growing period in this experiment was 12 weeks for the native and 10 weeks for the cultivated pasture.

Additional investigations on the relationship between water solubility and in vivo digestibility are recommended, to verify the results of this investigation.

Key Words: Brachiaria ruziziensis, chemical composition, in-vitro digestibility, native pasture, nutritive value, pasture quality

Introduction

Forage yield and nutritional qualities of pasture are influenced by numerous factors representing ecological conditions and management activities. Those factors include frequency of cutting, species composition, stage of maturity of plants, climatic conditions, soil fertility status and season of harvesting. As pasture gets mature it is characterised by high content of fibre with a higher grade of lignification and low protein content. Changes of quality during the growing period of grasses are particularly high under tropical climatic conditions due to the physiological differences to the temperate grasses (C4 vs. C3 grasses) and the higher photosynthesis rate at the increased radiation intensity in the tropics (Nelson and Moser 1994).

Under normal circumstances, cultivated pastures result in a remarkable higher yield and nutrient content relative to the native pastures. Several attempts have been made mainly in temperate regions to predict the digestibility of their forages by chemical, in situ and in vitro methods. The in vitro dry matter disappearance (Tilley and Terry 1963), enzymatic methods using cellulase (Donefer 1963, De Boever et al 1986, 1988; Kuhla and Weißbach 1993; Kuhla et al 1994; Weißbach et al 1999), the in vitro gas production test (Steingass and Menke 1986) and the nylon bag method (Ørskov and McDonald 1979) are alternative methods to estimate the nutritive value of forages in temperate regions. However their application on tropical grasses is not sufficiently tested.

The objective of this investigation was

• to assess nutrient yield in Hyparrhenia dominated native and cultivated (Brachiaria ruziziensis Germain and Evrard) pastures at three stages of harvest.

• to compare different chemical, in situ and in vitro methods and their relationships for estimating pasture quality.

Materials and methods

This investigation was carried out at the Wakwa research centre (Institute of Agricultural Research for Development, Wakwa) in Cameroon. Wakwa is located at an altitude of 1200 m above sea level. The plateau has a mild highland subtropical climate with a mean annual temperature of 22˚C, a relative humidity of 40 - 60% and a precipitation of 1600 - 1700 mm per annum falling mainly between April and October.

Pasture management

The treatments were three harvesting intervals at 8, 10 and 12 weeks arranged in a completely randomised block design in 6 replications separately for the cultivated and native pastures for two growing seasons during 1995 and 1996 (mid August - early November). Before the actual data collection of the experiment started the pastures were grazed at a stocking rate of 1.5 TLU/ha for 7 months. Then the pasture was managed without any application of input such as fertilizer.

Chemical analysis and in vitro and in situ measurements

From harvested pastures a representative sample of two kg fresh forage was taken, oven dried at 60°C for 48h hours and milled (1mm and 2 mm sieve size) for the different analyses. The chemical and the enzymatic cellulase analysis were done at the Institute of Animal Sciences of the Humboldt-University of Berlin, Germany. Chemical analysis consisted of proximate and detergent analysis (Naumann and Bassler 1997; Goering and van Soest 1970). The pepsin cellulase method was done according to De Boever et al (1986, 1988) and involved a pre incubation time in water at 80º C for 45 minutes before the addition of cellulase "Onuzuka R-10" from the yeast Trichoderma reesei. (Naumann and Bassler 1997).

Nylon bag degradation studies were carried out at Wakwa using two 18 month old steers (average weight 250 ± 15.0 kg). The animals were fed on a 1:1 mixture of native and Brachiaria hay as a basal diet, supplemented with 2.5 kg/day cotton seed cake (44% CP) and provided with mineral/vitamin pre mix and water ad libitum. The nylon bags were introduced by using sequential addition method (Osuji et al 1994) and incubated for 12, 24, 48 and 72 hours. The water solubility was estimated by soaking the bags in 39 º C water for 2 hours. Ten cm wide rubber fistula from Bar Diamond Inc. of Parma, Idaho, USA were used. The nylon bags (internal diameter 15 x 10 cm, with 53 µm pore size) were filled with 3 g of sample, corresponding to 11 mg/cm². Four replicates per treatment were incubated.

The potential degradable part of grasses (PD) was compared with results of Cellulase method, chemical analysis and the washing loss of nylon bags. Regression lines and correlation coefficients were calculated to prove the relationship between PD-values and the other values looking for alternative methods of quality estimation.

Calculations

The potential DM disappearence and the effective degradability of the forages using the nylon bags were described by using the model of Ørskov and McDonald (1979) as follows:

Y = a + b (1 - e ^-ct )

ED = a + bc /(k+c)

where:

Y = the potential disappearance of DM at time (t),
a = the rapid soluble fraction,
b = potentially degradable fraction and
c = rate of degradation of b.
k = passage rate at outflow rate of 3%/hour.

The potential degradability PD = a + b

The percentage organic matter solubility in cellulase (ELOS)), the cellulose digestibility of the organic matter (CDOM) and the insoluble organic matter of cellulase method (EuLOS) were calculated as follows:

ELOS (%) = % DM - % Ash - % Loss upon ashing

CDOM (%) = OMc x 10² / (100 - % Ash)

EuLOS = 1000 - Ash (g7kgDM) - (ELOS% x 10)

Data were analyzed using the general linear models (GLM) procedure of SAS (SAS 1991). Regression analysis for the inter-relationships among the variables were done using CORR procedure. Tests of significance were done using the Type III sums of squares.

The statistical model was:

Y_ijkl = µ + A_i + B_j + C_k +D_l + (A*B)_ij + (B*C)_jk + ε_ijkl

Where:

Y is observed variable_,
µ = overall mean;
A_i = fixed effect of year i;
B_j = fixed effect of pasture type j;
C_k = fixed effect of growing period k;
D_l = fixed effect of block k;
ε_ijkl = the experimental error.

Results and Discussion

Forage yield and nutrient content

The average forage yield was 2017 kg/ha, and significantly affected by the pasture type, length of growing period and year (Table 1).  The dry matter yield at 12 weeks regrowth was significantly (P<0.05) higher by 24 % compared to 8 weeks. The forage yield difference between Brachiaria and the native pasture was expected but much lower than described in the literature (Barnes 1996; Barsuala and Le Joly 1989; Ezenwa et al 1996; FAO 1967-1975). These authors reported that the cultivated Brachiaria pasture was usually fertilised and weeded against the invading species. The increase in forage yield in this experiment is not directly comparable with these results due to the length of harvesting interval (8-12 weeks) and no additional management practices apart from grazing before starting the experiment. Nevertheless significant differences in the forage yield between the two pasture types were observed in this experiment.

The average CP and NDF contents of the pastures were 5.3 and 68.2% respectively. Pasture type and regrowth lengths significantly influenced the nutrient content of the pastures. The native pasture had a lower protein content and a higher fibre content than Brachiaria. Among the different estimated fibre fractions of the pasture types, the strongest differences were found on the ADL, with a 30% higher content in native than in Brachiaria pasture.

Table 1 Forage yield (kg dry matter/ha) and nutrient content as affected by year, pasture type and length of growing period
Variables	n	Forage yield	Nutrient content, g/kg dry matter
			CP	CF	NDF	ADF	ADL
Year	72	***	ns	**	ns	ns	*
1995		2105	53.2	326	680	381	52
1996		1929	52.8	334	684	386	54
Pasture Type	72	*	*	*	***	***	***
Native	36	1926	48	358	720	419	60
Brachiaria	36	2108	58	302	644	347	46
Length of growing period	72	*	*	*	***	***	***
8 weeks	24	1798	59	322	669	374	52
10 weeks	24	2021	53	328	680	382	53
12 weeks	24	2232	48	340	700	394	54
R2		0.89	0.71	0.88	0.90	0.86	0.74
CV (%)		4.35	10.27	3.80	2.24	4.37	8.76
Mean ± SEM	72	2017 ± 87.8	53 ± 5.5	329 ± 12.5	682± 9.0	383± 16.7	53 ± 4.6
n = number of observations; CP = crude protein CF = crude fibre;   NDF = neutral detergent fibre; ADF = acid detergent fibre; ADL = acid detergent lignin; *** p < 0001; ** p < 001; * p < 005; ns  not significant

As the harvesting interval was delayed from 8 to 12 weeks, the CP declined by 23%, while the fibre fractions (CF, NDF, ADF) and ADL increased by 20 and 4% respectively. The lower quality obtained for both the pasture types in this study compared to previous reports at the Wakwa station (Piot and Rippstein 1975; Rippstein 1985) was a result of the low input management practices in this experiment, which represented the normal practice of the farmers at Adamawa plateau. The nutrient content of Brachiaria in this experiment, compared with literature results from other regions (FAO 1978; Moore and Miller 1988; Narmsilee et al 2002) showed a lower CP content by 23 to 60% but a similar content of CF and NDF while the ADL and ADF content was lower. This could be explained by the lack of N-fertilizer (regarding the CP content) and the shorter growing period of 8-12 weeks (regarding the fibre content). The decline of the quality results primarily from a change in the leaf to stem ratio (Ugherughe 1986) and the differences in quality between stems and leaves. The leaf content of switchgrass was changed from 47% to 26% between early and late stage of growth (Twidwell et al 1988). Additionally the leaf to stem ratio of grasses was much lower when grown in warmer than cold seasons (Deinum and Dirvin 1975).

The CF, ADF, and ADL content of Brachiaria was comparable with the late stage of temperate grasses under low management practices (Meak 2002) but it had 20% higher NDF and hemicellulose. The fibre and all estimated fractions in the native pasture were higher by 15-22% than the temperate grasses. The tendency of overproportional increases of the hemicellulose content in warm season grasses is also described by Buxton and Fales (1994) and Van Soest (1982).

In situ and in vitro digestibility

The results of digestibility by cellulase solubility method showed that Brachiaria pasture had 40% higher ELOS and CDOM, while the digestibility estimated by nylon bag degradability techniques was only 7% higher, relative to the native pasture (Table 2).

Table 2. Measurements by nylon bag degradability and cellulase solubility techniques of Brachiaria and native pastures harvested at different length of growing period during 1995 and 1996 (% on dry matter basis)
Variables	n	a (0h)	PD = (a+b)	ED	ELOS	CDOM	EuLOS
Year	36	**	ns	ns	ns	ns	ns
1995	18	19	54	41	38	41	54
1996	18	18	52	40	39	42	53
SEM		00.3	0.6	0.3	1.6	0.4	0.6
Pasture Type	36	***	***	***	***	***	***
Native	18	16	51	40	32	32	55
Brachiaria	18	21	55	42	45	45	46
SEM		00.3	0.6	0.3	1.6	1.6	0.6
Growing period	36	***	***	***	*	ns	*
8 weeks	12	22c	56b	44c	39a	43	52a
10 weeks	12	19b	54ab	41b	.38b	41	54b
12 weeks	12	16a	49a	37a	38b	41	54b
SEM		0.4	0.8	0.4	02.0	0.8	0.7
R2		0.80	0.74	0.76	0.91	0.89	0.90
mean ±		19	53	41	38	42	53
SEM		0.2	0.3	0.2	0.3	0.3	0.3
n = number of observations; a= intercept of nylon bag method; PD = potential degradability of DM,  ED = effective degradability at outflow rate 3%/hour; ELOS = cellulase solubility of the organic matter; CDOM = cellulase digestibility of organic matter; EuLOS = insoluble organic matter in cellulase. Figures with different letters within columns indicate significance values *** p < 0001; ** p < 001; * p < 005; ns not significant

Growing period did not affect the cellulose solubility significantly, however, a strong negative correlation was obtained between the length of growing period and the nylon bag degradability.

Correlations

The nylon bag degradability was determined with local cattle in Cameroon and the PD values were calculated according Ørskov and McDonald (1979) and were taken as the reference for assessing the digestibility values obtained by other methods in this investigation (Table 3).  The coefficients of determination (R²) between the different chemical parameters and the PD values were between 0.11 (ADL) and 0.31 (NDF and CF). This low level of correlation agrees with results of investigations in temperate regions, were strong correlation exists between fibre content and digestibility only for pastures with high management activities (application of fertilizer, 3-4 cuts per year) (DLG 1997). Unlike grass-legume mixed natural pastures (mostly clovers), relatively strong correlations for pure grass pastures has been reported by Jones (1970), Burrit et al (1984, 1985), Van Soest (1982), Ford (1978), Fales et al (1991) and Smith et al (1972). The present result of higher correlation trend between digestibility and CF and NDF content unlike the ADL, confirms the results of Südekum et al (1990) and Van Soest (1982). From this experiment it can be concluded, that parameters of chemical analysis are not qualified to predict the digestibility of tropical pastures.

Table 3 Linear function of pasture: The relation between the PD value and chemical, in situ and in-vitro parameters adjusted
Linear function y = PD =	adjusted R2	sE	sign.	n
91.06 - 0.056 * NDF	0.31	0.09	***	72
73.95 – 0.055 * ADF	0.25	0.10	***	72
63.59 – 0.199 * ADL	0.11	0.55	**	72
78.16 – 0.710 * CF	0.31	0.12	***	72
37.79 + 0.775 * ws	0.78	0.06	***	72
12.54 + 0.991 * ED	0.92	0.04	***	72
42.11 + 0.282 * ELOS	0.18	0.06	***	68
40.63 + 0.295 * CDOM	0.23	0.06	***	68
68.19 – 0.029 * EuLOS	0.19	0.01	***	68
ws= water solubility from nylon bag; PD = potential degradability of DM; ED = effective degradability at outflow rate 3%/hour; ELOS = cellulase solubility of the organic matter; CDOM = cellulase digestibility of organic matter; EuLOS = insoluble organic matter in cellulase. R2 = coefficient of determination, n = number of observations

The correlation between the PD values and the cellulose solubility in this experiment was found to be low. The coefficients of determination varied between 0.18 and 0.23, which indicates that the latter is not an effective method to be used as a reference. Maybe the method must be adjusted to tropical forages in line with the lower and slower degradability of the fibre fractions. A comparison of different methods to predict in vivo dry matter digestibility (Navarante et al 1990) showed the highest error of estimation for the enzymatic cellulose method in C4 grasses. Other similar works indicate that the type of cellulase enzyme used for solubility and the species of the forage to be tested determine the level of cellulase digestibility (Gabrielson 1986; DeBoever et al 1988). Weiss (1995) postulated that the structure of the cell walls is responsible for the degradation of organic matter by cellulase.

Estimation of DM water solubility from the nylon bag had a high positive correlation coefficient (R² = 0.72) with the potential degradability of DM (Figure 1). This suggests that estimation of water solubility is a potential means of prediction degradability. In addition to it's high precision, it is fast, cheap and easy for application. This agrees with the results of Ly and Preston (1997, 2001) and Ly et al (1997) who determined water solubility by washing the nylon bags for 90 minutes in a commercial washing machine. The correlation between in sacco dry matter loss and washing dry matter loss increased with prolonged washing time from 30 to 120 minutes (Nguyen Van Lai and Nguyen Thi Thu Huong (1999).

Figure 1. Correlation between potential degradability in the rumen after
72 hours incubation and water solubility

The potential degradable forage yield of the pastures harvested at the different lengths of growing period did not vary significantly, despite the trend of increased PD forage yields of the native pasture as harvesting periods got longer during the experimental period (Figure 2). The PD forage yield for Brachiaria appeared to be highest at the 10 weeks harvesting stage.

Figure 2. Potentially degradable dry matter yield of Brachiaria and native pastures
harvested after different periods of re-growth

Conclusions

Acknowledgements

The authors gratefully acknowledge the financial support of the Deutscher Akademischer Austausch Dienst (DAAD) as well as the staff of the Wakwa research institute for assistance with the trial.

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