Wool growth in Merino wethers fed lupins untreated or
treated with heat or formaldehyde, with and without a
supplementation of rumen protected methionine
M. Rodehutscord
*, P. Young, N. Phillips, C.L. White
CSIRO Division of Animal Production, Private Bag, PO Wembley, Western Australia 6014, Australia
Received 2 June 1999; received in revised form 9 August 1999; accepted 28 August 1999
Abstract
Lupins were treated by either heat (1158C for 1 h) or by formaldehyde (0.4 g 100 gÿ1 crude protein). The fractional rate of disappearance of N from dacron bags suspended in the rumen of steers was reduced by either treatment. Assuming a rumen outflow rate of 0.03 hÿ1, effective rumen
degradability was 0.96, 0.94 and 0.92 for protein and 0.84, 0.82 and 0.81 for dry matter in untreated, heat- and formaldehyde-treated lupins. Concentration of acid detergent insoluble nitrogen in lupins was not enhanced by treatment.
A 32 factorial experiment was performed with Merino wethers to study the effect of treating lupins on wool growth and body weight gain on a feeding level slightly above maintenance. Factors were lupin treatment (untreated, heat- and formaldehyde-treated) and supplementation of a rumen protected methionine (3 g methionine per day, yes or no). The diet contained (kgÿ1) 620 g chaffed oaten hay, 350 g broken lupins and 30 g mineral premix and was supplied once daily to sheep at a level of 900 g per day air dry or approximately 8 MJ per day ME. Seventy-two sheep were fed the diet containing untreated lupins without methionine supplementation during a 4 week pretreatment period and were subsequently allocated to one out of the six treatments according to pretreatment wool growth rate (n12 sheep per treatment). Treatment lasted for 8 weeks. Comparative clean wool growth rate was determined on mid-side patches of approximately 100 cm2shorn in 4
week-intervals. Faeces and urine were collected for 7 days at the end of the experiment with six sheep per treatment. Both body weight gain and clean wool growth were not significantly effected by either treatment of lupins. Supplementation of rumen protected methionine significantly increased both body weight gain (by 27%) and clean wool growth. The effect of supplementary rumen protected methionine on clean wool growth was twice as high in sheep fed either heat- or
formaldehyde-82 (1999) 213±226
*Corresponding author. Present address: Institut fuÈr ErnaÈhrungswissenschaften, UniversitaÈt Halle-Wittenberg,
06099 Halle, Germany. Tel.:49-345-5522701; fax:49-345-5527124
E-mail address: rodehutscord@mluters1.landw.uni-halle.de (M. Rodehutscord)
treated lupins (37 and 36%, respectively) as compared to sheep fed untreated lupins (19%). Sulphur, but not nitrogen concentration in clean wool was significantly increased by supplementation of rumen protected methionine. The efficiency of utilisation of metabolisable protein (defined according to [AFRC, 1993. Energy and protein requirements of ruminants. CAB International, Wallingford]) for N retention was improved by 13, 22 and 27% for diets containing untreated, heat-and formaldehyde-treated lupins due to supplementary rumen protected methionine. Organic matter digestibility and daily faecal N excretion were unaffected by lupin treatment and by supplementation of rumen protected methionine, indicating an equal ME supply to all sheep. Correspondingly, the ratio of total purine derivatives to creatinine in urine was not significantly affected by either experimental factor. 79% of total N in urine was present as urea.
It is concluded, that treating lupins with formaldehyde or heat cannot be recommended as a means to improve the lupin protein quality for wool production unless extra rumen protected methionine is supplemented.#1999 Elsevier Science B.V. All rights reserved.
Keywords: Wool growth; Sheep; Lupins; Protein; Protection; Protected methionine
1. Introduction
During the dry summer and autumn periods in the West-Australian Mediterranean climate zone pasture growth completely stops and wool growth of sheep as well as wool quality are reduced, combined with a loss in body weight (BW) of sheep. Supplementary grain feeding is commonly practised during this period with the Australian sweet lupin
seed (Lupinus angustifolius) as a grain preferred by farmers. It is available on many
West-Australian farms because it forms part of the rotation. Lupins are easy to handle and to feed (spread on the ground) and they are high in metabolisable energy (ME) concentration.
Wool growth is a function mainly of the amount of amino acids reaching the intestine (Hynd and Allden, 1985) rather than energy supply (Black et al., 1973; Reis et al., 1992). Furthermore, the amino acid pattern of the protein which reaches the intestine may effect wool growth since sulphur-containing amino acids (SAA) are first limiting in terms of wool protein synthesis (Reis and Tunks, 1978). Therefore, the ruminally degradable proportion of dietary protein becomes a critical factor for wool growth and, on isonitrogenous diets, wool growth responds well to less degradable proteins, particularly when they are high in SAA concentration (Coombe, 1992; Masters and Mata, 1996; White et al., 1999). Consequently, SAA supplementation in the form of methionine, cysteine or cystine is effective for enhancing wool growth if degradation in the rumen is avoided (Reis and Tunks, 1978).
Lupins contain approximately 32±35% crude protein (Petterson et al., 1997), which makes them appear to be an effective protein supplement for sheep. But lupin protein is highly degradable in the rumen (Antoniewicz et al., 1992; AFRC, 1993; White et al.,
1999) and contains only, on an average, 2.2 g methioninecystine 100 gÿ1
CP (Degussa, 1996). This, in fact, makes lupins a low quality protein supplement in terms of wool growth, and a better wool growth response than with lupins could be detected with less degradable proteins higher in SAA concentration such as canola meal (Masters and Mata, 1996). Surprisingly, even supplementary oats prove equal to or better than lupins in
promoting wool growth (White et al., 1999). As there obviously are some advantages from using lupins as a supplement, we were aiming at trying to improve the value of lupin protein for wool production.
Both formaldehyde treatment (Crooker et al., 1986; Antoniewicz et al., 1992) and heat treatment (Satter, 1986) were shown to be effective means to reduce the ruminal
degradation of protein in-vitro. Toasted broken lupins (3 min at 1328C) showed
remarkably lower in situ protein degradability as compared to untreated lupins without any difference in intestinal digestibility of rumen undegraded protein (Goelema et al., 1998). Wool growth in sheep fed a lupin based diet responded well to an increase in methionine supply by either genetic enhancement of methionine concentration in lupin protein (White et al., 1998) or by supplementation of free methionine protected against degradation in the rumen (Mata et al., 1995; White et al., 1999). It was the aim of the present study to investigate how sheep fed near maintenance on a lupin based diet respond, in wool growth and body weight gain (BWG), to treated lupin protein and to
rumen protected methionine. A 32 factorial experiment was conducted with the factors
`lupin treatment' (untreated, heat- or formaldehyde-treated) and `methionine supple-mentation' (yes or no).
2. Materials and methods
2.1. Lupin treatment
Lupins (L. angustifolius, mixed varieties) were taken from one batch. For heat
treatment, whole seed lupins were put in layers of approximately 2 cm high in drawers with a perforated stainless steel bottom. Drawers were put into a large-size conventional oven dryer and lupins remained inside for 1 h after air temperature had reached its
maximum of 1158C. Lupins were then removed from the oven and spread in a thin layer
on the floor in order to cool down immediately. For formaldehyde treatment, lupins were
gristed (course ground) and mixed with formaldehyde (0.4 g 100 gÿ1
CP; Hamilton et al., 1992). This level was chosen in order to avoid a reduction in intestinal digestibility of ruminally undegradable protein which potentially could result from a too high formaldehyde level.
Untreated and heat treated lupins were cracked in a hammer mill (1 cm hole diameter in the sieve) before being mixed into the diets. Analysed concentrations of dry matter and crude protein are shown in Table 1 for the three different lupin batches. Amino acid composition was determined for the untreated lupins only.
2.2. Diets
Diets consisted of chaffed oaten hay, lupins and a mineral premix (Siromin1
) (Table 2)
and were calculated to contain approximately 8 MJ ME kgÿ1
DM. Components were mixed in batches of 500 kg in advance for the entire experiment and stored in bags at room temperature until feeding. Diets for half of the animals were supplied with 10 g per
day Lactet1 (Nippon Soda), a product containing 30% DL-methionine, which can be
Table 1
Analytical data for lupins (results expressed on an air dry basis)
Lupins
Untreated Heat-treated Formaldehyde-treated
Dry matter (g kgÿ1) 906 943 911
Crude protein (g kgÿ1) 302 304 283
ADINa(g kgÿ1) 0.45 0.45 0.36
Amino acids(g/16 g N)
Alanine 3.5
Arginine 10.1
Aspartic acid 9.8
Cystine 1.45
Glutamic acid 19.0
Glycine 4.2
Histidine 2.6
Isoleucine 4.0
Leucine 6.6
Lysine 4.6
Methionine 0.6
Phenylalanine 4.0
Proline 4.0
Serine 4.8
Threonine 3.4
Tryptophan 0.8
Valine 3.9
aADIN: acid detergent insoluble nitrogen.
Table 2 Diets
Lupins Diet
Untreated Heat-treated Formaldehyde-treated
Composition(g kgÿ1)
Oaten hay 620 620 620
Lupins, untreated 350 ± ±
Lupins, heat-treated ± 350 ±
Lupins, formaldehyde-treated ± ± 350
Mineral mixa 30 30 30
Analysed
Dry matter (g kgÿ1) 921 927 923
Crude protein (g kgÿ1DM) 132 136 120
Sulphur (g kgÿ1DM) 3.1 3.6 2.7
aSiromin1contained (g kgÿ1mineral mix): Na 176, K 116, Ca 48, S 39, P 15, Mg 4, Fe 1.94, Zn 1.16, Mn
0.58, Cu 0.116, Mo 0.04, Co 0.08, Se 0.006, I 0.004, Ni 0.004, Cr 0.004, V 0.004, B 0.004.
assumed effectively protected against degradation in the rumen of sheep fed at maintenance level (Muramatsu et al., 1994; Mata et al., 1995). Lactet was not mixed into the diet but separately preweighed in small plastic vessels on a daily basis and spread over the feed immediately after this had been put into the troughs. Amount of Lactet was calculated to supply approximately twice the amount of absorbable SAA supplied with the untreated lupin diet.
2.3. Animals, feeding and sampling
Seventy-two Merino wether sheep weighing, on an average, 34 kg were selected from the Yalanbee research station flock and brought into the animal house where they were penned individually on wooden slatted floors. Sheep were approximately 5 months old and had been weaned at 3 months of age. They were given an adjustment period of 3 weeks during which the amount of feed was increased from 600 to 900 g per day (air dry). This quantity was calculated to allow for a ME supply slightly above maintenance and it was kept constant throughout the entire experiment. A 4 week pretreatment period followed for wool growth measurement under standardised feeding conditions. During adjustment and pretreatment all sheep were fed the diet containing untreated lupins without Lactet. Twelve sheep were allocated to one out of the six dietary treatments stratified according to BWG and wool growth during the pretreatment period. Dietary treatments lasted for 8 weeks. Diets (900 g per day air dry) were given in one meal in the morning. No feed refusals were recorded. Drinking water was continuously available from nipple drinkers.
Body weight was recorded at weekly intervals. A mid-side patch of approximately
100 cm2was shorn with small animal clippers (ANDIS with a model AG No. 40 cutting
head) and a dyeband was applied to wool alongside the patch at the beginning of the pretreatment period (Langlands and Wheeler, 1968). Sheep were shorn on these patches and dyebands were applied every 4 weeks (end of pretreatment, after 4 and 8 weeks on treatment). During the last 7 days on treatment, faeces were quantitatively collected from six sheep out of each treatment. Sheep were fitted with harnesses and faeces were collected in plastic bags as described by Cole et al. (1996). Pizzle harnesses were attached and urine was collected in bottles underneath the floor. Bottles contained 100 ml diluted sulphuric acid (10% v/v) to keep pH in bulked urine samples below 2. As leakage of the urine collection unit could not be completely avoided for all animals, urinary excretion cannot be regarded as quantitatively determined. Concentration of different urinary N fractions will, therefore, be expressed in relation to urinary creatinine concentration. However, collection of urine was complete for most of the animals and we estimate that the spot sample comprised at least 80% of total urine from 7 days for all animals.
2.4. Degradability study
In situ degradability was determined for untreated and treated lupins in four
rumen-cannulated steers (600 kg BW) fed on a maintenance diet which consisted of 50%
cereal hay, 24% lupins, 24% faba beans and 2% mineral mix (Siromin1). Lupin samples
were ground in a mill with a 3 mm sieve and weighed into dacron bags with a 50mm pore
size. Lines with the bags were washed for one wash and spin cycle in an automatic washing machine (16 min) and then suspended in the rumen of the steers for 48 h at a maximum. One set of bags was dried and weighed after removal from the washing machine to give the zero time solubility, and the remainder were removed from the rumen at intervals of 2, 4, 8, 12, 16, 24 and 48 h (one bag per steer per lupin batch per time). Bags with the residual material were immediately placed in cold water, and then washed
for one cycle in the automatic washing machine, dried at 558C for 24 h and analysed for
nitrogen. The exponential functionyab(1ÿeÿct) was fitted to the data, whereyis
the cumulative disappearance of nitrogen from the bags depending ont,tthe incubation
time (hours),athe soluble fraction (determined as time zero washout),bthe potentially
degradable fraction and cthe fractional rate of degradation of feed nitrogen per hour
(AFRC, 1993).
2.5. Analysis
Greasy wool samples were washed for 6 min in hot water (658C) and Scour (1 ml/l
water) (Dolmar Australia), a commercially available heavy duty wool scouring compound and dried in a humidified room for 72 h before weighing. Results for clean wool growth (CWG) are expressed on a 88% dry matter basis (humidified clean wool). Clean wool growth during treatment is covariance adjusted for individuals based on
respective pretreatment values. Feed and faeces were dried at 708C and ground. Ash was
determined after ignition in a muffle furnace at 5508C. Feed, faeces and urine were
analysed for Kjeldahl-N (Tecator-system) and crude protein was calculated as N6.25.
Urea-N was analysed on a Cobas Mira clinical chemistry analyser (Roche1
) using a
commercially available kit (Sigma Diagnostics1
). Purine derivatives in urine were determined according to Chen and Gomes (1992). Wool nitrogen and sulphur were
determined on a Leco1
NS2000 nitrogen and sulphur analyser using total combustion method. Statistical analysis and parameter estimate were performed on a personal computer using the software package SPSS 7.5 for Windows.
3. Results
Supplementation of rumen protected methionine from Lactet significantly increased BWG of sheep by approximately 15 g per day or 27% without a significant interaction with the lupin treatment (Table 3). Body weight gain of sheep receiving either of the treated lupins was numerically lower than in sheep fed the untreated lupins but this effect was statistically not significant. Clean wool growth during pretreatment averaged
0.870.02 mg cmÿ2
per day (meanSEM). In both 4 week treatment periods,
supplementation of rumen protected methionine significantly improved CWG by, on
average, 0.12 and 0.26 mg cmÿ2
per day in weeks 1±4 and weeks 5±8, respectively (Table 3). This effect of rumen protected methionine was higher when treated lupins rather than untreated lupins were fed, particularly in weeks 5±8, when the interaction reached
significance level (P0.058). To confirm that this interaction was statistically valid, the
wool data for weeks 5±8 lupin treatments were partitioned into two contrasts of 1 d.f.
each, Ð untreated versus treated and heat treated versus formaldehyde treated. Virtually all the variance was associated with the comparison of no treatment versus treatment and
its interaction with methionine (P0.017). This clearly shows that there was an
interaction between grain treatment and methionine, and that heat was equivalent to formaldehyde in treating lupins. An inspection of the data shows that the results for period 1 were in the same direction as for period 2, just not significant using the original analysis. This difference between periods is often the case for wool responses because of the lag phase.
Treating lupins by either heat or formaldehyde appeared to reduce CWG when no rumen protected methionine was supplemented but this effect of lupin treatment was not significant in either period. The `yield', which is commonly understood as the proportion of clean wool in greasy wool, was, on an average, 80 and 77% in weeks 1±4 and weeks 5±
8, respectively. In weeks 5±8, rumen protected methionine (P0.064) but not lupin
treatment (P0.346) improved yield (Table 3).
Pretreatment concentrations of nitrogen and sulphur in clean wool were 18.20.31
and 3.170.04 g 100 gÿ1
, respectively. There was a tendency towards a higher concentration of both nitrogen and sulphur in wool due to heat treatment of lupins in weeks 1±4, but lupin treatment did not significantly affect nitrogen and sulphur concentration of wool grown in weeks 5±8 (Table 4). Supplementation of rumen protected methionine did not affect wool nitrogen but significantly improved wool sulphur concentration in both periods.
Neither lupin treatment nor rumen protected methionine supplementation significantly affected organic matter digestibility (OMD), but OMD was numerically lower in the diet containing formaldehyde-treated lupins as compared to the other diets (Table 5). Daily
faecal N excretion was, on an overall average, 4.360.07 g per day without an effect of
either factor.
The concentration of urinary total N in relation to creatinine was not affected by lupin treatment or methionine supplementation but the concentration of urea N was lower
Table 3
Body weight gain (BWG) and clean wool growth (CWG) in sheep fed differently treated lupins without or with a rumen protected methionine (Lactet). Values for BWG and CWG are covariance-adjusted LS means (n12 sheep per treatment)
Lupin treatment
None Heat Formaldehyde Pooled P-value (ANOVA)
Lactet ÿ ÿ ÿ SE Treatment Lactet Treatment
Lactet
BWG (g per day) 60 73 56 71 51 69 2.3 0.353 <0.001 0.801
CWG(mg cmÿ2per day)
Weeks 1±4 0.98 1.06 0.91 1.03 0.94 1.11 0.02 0.129 <0.001 0.244 Weeks 5±8 0.90 1.07 0.83 1.14 0.86 1.17 0.02 0.571 <0.001 0.058
CWG(% of greasy wool)
Weeks 1±4 80 82 79 77 81 80 0.8 0.361 0.990 0.539 Weeks 5±8 74 76 76 79 75 79 0.7 0.346 0.064 0.924
Table 4
Nitrogen and sulphur concentration (g 100 gÿ1) in clean wool of sheep fed differently treated lupins without or
with a rumen protected methionine (Lactet) (n12 sheep per treatment)
Lupin treatment
None Heat Formaldehyde Pooled P-value (ANOVA)
Lactet ÿ ÿ ÿ SE Treatment Lactet Treatment
Lactet
Nitrogena
Weeks 1±4 16.9 16.7 17.5 18.0 16.9 17.7 0.14 0.020 0.174 0.264 Weeks 5±8 17.7 17.1 17.4 17.0 17.6 17.5 0.11 0.351 0.092 0.628
Sulphurb
Weeks 1±4 3.18 3.52 3.27 3.76 3.11 3.51 0.04 0.014 <0.001 0.592 Weeks 5±8 3.33 3.74 3.26 3.71 3.27 3.58 0.04 0.291 <0.001 0.603
aNitrogen concentration during pretreatment period was 18.20.31 g 100 gÿ1. bSulphur concentration during pretreatment period was 3.170.04 g 100 gÿ1.
Table 5
Organic matter digestibility (OMD) and faecal N excretion in sheep fed differently treated lupins without or with a rumen protected methionine (Lactet) (n6 per treatment)
Lupin treatment
None Heat Formaldehyde Pooled P-value (ANOVA)
Lactet ÿ ÿ ÿ SE Treatment Lactet Treatment
Lactet
OMD (%) 73.8 72.4 73.1 73.2 71.8 71.9 0.3 0.207 0.512 0.585 Faecal N (g per day) 4.43 4.48 4.56 4.21 4.21 4.30 0.07 0.501 0.613 0.367
Table 6
Urinary excretion of total N and nitrogen fractions in sheep fed differently treated lupins without or with a rumen protected methionine (Lactet) (n6 sheep per treatment), expressed in relation to creatinine excretion (mgÿ1creatinine). Mean creatinine concentration was 11.1 mg lÿ1urine
Lupin treatment
None Heat Formaldehyde Pooled P-value (ANOVA)
Lactet ÿ ÿ ÿ SE Treatment Lactet Treatment
Lactet
Total N (mg) 13.9 12.4 14.1 13.5 12.4 12.6 0.4 0.345 0.368 0.600 Urea N (mg) 10.7 10.0 11.3 10.9 10.5 9.5 0.2 0.109 0.100 0.866 Allantoin (mmol) 7.1 4.1 3.7 4.2 4.2 5.5 0.5 0.316 0.647 0.121 Xanthine
hypoxanthine (mmol)
2.2 1.7 1.6 1.6 1.8 1.4 0.1 0.037 0.017 0.243
Uric acid (mmol) 2.0 1.7 2.2 1.6 1.8 1.7 0.1 0.935 0.102 0.628 Total purine
derivatives (mmol)
11.4 7.6 7.5 7.4 7.9 8.6 0.5 0.153 0.209 0.068
(P0.100) in sheep which received the methionine supplemented diets than in sheep fed the unsupplemented diets (Table 6). Concentration of total purine derivatives was not
significantly influenced but the concentration of xanthinehypoxanthine was reduced
both by lupin treatment (P0.037) and by methionine supplementation (P0.017).
Methionine supplementation also tended to reduce uric acid concentration (P0.102).
Allantoin concentration was not significantly affected by either factor.
Treatment of lupins did not alter the potential degradability (ab) of dry matter and
protein. However, the rate of degradationc, particularly for protein, was affected by lupin
treatment (Table 7 and Fig. 1).
4. Discussion
Wool growth is mainly limited by the amount of amino acids reaching the intestine, estimated for example as digestible protein leaving the stomach (DPLS; Freer et al.,
Table 7
Parametera,bandcestimated for the functionyab(1ÿeÿct) to describe the cumulative disappearance of lupin dry matter and nitrogen from dacron bags suspended in the rumen of steers for 48 h at a maximum
a b c R2 syx
Dry matter
Untreated 0.32 0.66 0.11 0.93 0.05
Heat-treated 0.28 0.70 0.10 0.96 0.05
Formaldehyde-treated 0.29 0.71 0.08 0.95 0.05
Nitrogen
Untreated 0.38 0.61 0.64 0.98 1.94
Heat-treated 0.35 0.64 0.38 0.98 2.20
Formaldehyde-treated 0.30 0.68 0.30 0.94 4.18
Fig. 1. Disappearance of lupin N from dacron bags suspended into the rumen of steers. The course of disappearance was described by the functionyab(1ÿeÿct) and estimated parameter are listed in Table 7. Data shown are for the first 24 h of incubation only.
1997) or metabolisable protein (MP AFRC, 1993). Concentration of SAA in wool is
approximately 11 g 100 gÿ1
total amino acids, mainly present as cysteine, whereas it is
around only 3.5 g 16 gÿ1
N in total microbial amino acids, largely present as methionine (Storm and Orskov, 1984), making SAA first limiting for wool growth under most conditions, particularly at low ME supply. In the present experiment, pretreatment of lupins did not affect the potential protein degradability but did reduce the rate of protein degradation determined in situ in steers (Table 7). Using the parameter shown in Table 7,
the supply of MP to sheep, based on AFRC (1993) and a rumen outflow rate of 0.03 hÿ1,
was approximately 44 g per day when untreated lupins were fed. MP supply increased by 5 and 11% when heat and formaldehyde, respectively, were applied to lupins.
No effect of either treatment, however, could be detected in growth and sulphur concentration of clean wool in the present study in the Lactet-free diets. This raises some doubt about whether the theory about the correlation between wool growth and intestinal MP supply holds true in general or whether it needs modification with regard to the SAA-concentration in MP. There is an indication from amino acid analysis of residues after rumen incubation that the degree of degradation is not equal for all amino acids (Crooker et al., 1986; SuÈdekum and Andree, 1997; van Straalen et al., 1997; Chiou et al., 1999) and that it could be higher for methionine than for crude protein (Crooker et al., 1986). Furthermore, Ashes et al. (1984) reported a lower availability of labelled cysteine from casein in sheep as formaldehyde concentration increased. Thus, protection of protein does not necessarily mean an equal protection of SAA and a proportional increase in intestinal SAA flow. Additionally, when intermediary supply of methionine is increased to a lower degree than the supply of other essential amino acids, methionine oxidation might be increased due to a generally increased oxidation of amino acids and, consequently, methionine is no longer available to the wool follicles. Sulphur concentration in wool positively responded to methionine supplied from Lactet but not to lupin treatment, which can be taken as a further indication that intestinal SAA supply was not improved by treating lupins. Therefore, no effect on wool growth can be expected even when a higher level of protection of lupin protein is achieved with a higher dose of formaldehyde or a higher exposure of lupins to heat. On the other hand, a higher response to treatment than in lupins can be expected with proteins which are higher in SAA concentration.
Sheep tended towards a reduced wool growth when fed treated lupins compared to untreated in both periods (weeks 1±4 and 5±8, respectively), but response to supplementation of protected methionine was much stronger in weeks 5±8 than it was in weeks 1±4 of the experiment. Masters and Mata (1996) found that the positive effect of feeding canola meal instead of lupins to ewes continued after completion of the experiment when sheep had already been returned to the paddock for 3 weeks. A lag phase in the response to a change in diet is indicated also by other studies and could be caused both by the time required for the microbial population in the rumen to adjust and by the wool follicles to adapt to a change in methionine supply. Therefore, in discussing long-term effects of lupin treatment and methionine supplementation, weeks 5±8 appear to deliver more generally applicable results than weeks 1±4.
In weeks 5±8, the effect of methionine supplementation, on CWG, was different
depending on the lupin treatment (P0.058) (Table 3). While methionine
supplementa-tion increased wool growth by 19% when the untreated lupins were fed, it improved wool
growth by 37 and 36% when heat- or formaldehyde-treated lupins were fed. There is no indication from the purine : creatinine-ratio in urine that methionine supplementation positively affected microbial protein synthesis in the rumen, as reported in earlier experiments (White et al., 1999). Amino acids other than methionine therefore must have limited wool growth in the Lactet-supplemented sheep fed untreated lupins. Increased incorporation of sulphur from rumen protected methionine into wool with pretreated lupins could consequently happen only because the supply of these potentially limiting amino acids was improved. This is further strong evidence that there was an increased flow of MP due to heat or formaldehyde treatment in the present experiment. For practical application, however, none of the treatments tested here can be recommended to improve the protein quality of lupin grain for sheep, unless a cheap enough source of rumen protected methionine is available to the farmer. If methionine is applied, its value for wool growth can be doubled by pretreating lupins with heat or formaldehyde.
Organic matter digestibility was not significantly effected by treatment of lupins (Table 5). This can be interpreted as a strong indication for an equal ME supply to all groups of sheep in this experiment and corresponds to the unaffected purine excretion. However, there was a tendency towards a reduced OM digestibility due to formaldehyde treatment and this conforms with a lower (not significantly) BWG when formaldehyde treated lupins were fed as compared to the other diets (Table 3). In situ data show that the rate of degradation was lower in formaldehyde-treated than in untreated lupins (Table 7) and the
effective dry matter degradability, calculated for a rumen outflow rate of 0.03 hÿ1, was
0.84 and 0.81 for untreated and formaldehyde-treated lupins, respectively. This indicates a reduced fermentation in the rumen which could explain the lower OMD and the lower BWG of sheep fed the formaldehyde-treated lupins. Body weight gain was improved by rumen protected methionine in a similar matter as wool growth, indicating that SAA supply limited protein retention in fleece-free body as well as in wool. This is in agreement with results from Mata et al. (1995) and shows that weight gain is not simply a function of ME and MP supply but that it depends on amino acid pattern of MP as well.
The level of formaldehyde used to treat lupins (0.4 g 100 gÿ1
CP) was low and chosen to avoid a depression in intestinal digestibility that could result from an overprotection of the protein. Different levels of formaldehyde have been tested for their effect on in situ
degradation of protein from lupins (1.0±4.0 g 100 gÿ1
CP, Antoniewicz et al., 1992) and
soybean meal (0.6±1.8 g 100 gÿ1
CP, Crooker et al., 1986). In both cases, a remarkable reduction in in situ protein degradation was achieved on the lowest formaldehyde level with only a minor effect of a further increased formaldehyde level. The effect on protein
degradation found in the present experiment using a formaldehyde level of 0.4 g 100 gÿ1
CP was less and it appears possible that a higher level of protection would have been achieved with a higher formaldehyde dose. However, for reasons mentioned above it is unlikely that this could improve wool growth. A negative effect on intestinal digestibility of undegraded protein is unlikely as indicated by ADIN concentrations (Table 1) and by the fact that a negative effect of formaldehyde treatment on intestinal digestibility of
crude protein from lupins was not obvious up to a formaldehyde level of 4.0 g 100 gÿ1
CP (Antoniewicz et al., 1992).
Pretreatment temperature applied to lupins in this study was comparatively low
(1158C) but pretreatment time was long (1 h). Regarding the level of protection achieved
it is generally accepted that total heat input (temperaturetime) is important rather than temperature alone (Satter, 1986). Goelema et al. (1998) more than doubled the
undegradable protein supply from lupins by pressure toasting (1328C for 3 min) without a
negative effect on intestinal digestibility of undegraded protein. This indicates that we did not achieve the optimum level of protection in the present study. However, a positive effect on wool growth is not necessarily to be expected from a higher level of protection due to the low SAA concentration in lupin protein as indicated above.
As calculated from BWG and wool growth data, sheep retained without methionine supplementation, on average, 2.7 g per day N (Table 8) while N intake during this experiment was, on an average, 17.2 g per day. This demonstrates the low overall N utilisation (16%) in low input wool production systems. Approximately 12 g per day N were excreted via urine only, 79% of which was present as urea which can be rapidly degraded to ammonia and carbon dioxide.
Current protein evaluation systems for ruminants work with constant efficiency figures, for example for the conversion of MP into product. AFRC (1993) gives a low fractional utilisation of MP for wool protein accretion of 0.26 while it is 0.59 for retention in fleece-free body. The SCA (1990) reports a mean gross fractional efficiency of MP for wool growth in Merino sheep of 0.12±0.18, or after maintenance MP is subtracted, 0.2±0.25. With certain assumptions, the utilisation of MP can be calculated for the present experiment in order to calculate the effect of a protected methionine supplementation on MP utilisation for protein retention as a whole (no partition between N retention in wool and body protein) (Table 8). In the present experiment, estimated MP supply to sheep ranged between 44 and 49 g per day, based on the equations of AFRC (1993) and the degradability parameter determined in steers. Total wool growth was calculated from dyeband results. Nitrogen retention in fleece-free BWG (g per day) was calculated as
0.16BWG (kg per day)(160.4ÿ1.22 BW (kg)0.0105 BW2 (kg)) (modified
Table 8
Calculation of the efficiency of utilisation of metabolisable protein (MP) depending on lupin treatment and methionine supply from Lactet
Lupin treatment None Heat Formaldehyde
Lactet
ÿ ÿ ÿ
MP supply (g per day) 44 44 46 46 49 49
BWG (g per day) 60 73 56 71 51 69
CWGa(g per day) 8.5 10.0 8.7 10.4 8.3 11.1
N accretion(g per day)
In fleece-free BWb 1.3 1.5 1.2 1.5 1.1 1.4
In woolc 1.5 1.7 1.5 1.8 1.5 1.9
Total 2.8 3.2 2.7 3.3 2.6 3.3
N accretion/N supplied from MP 0.40 0.45 0.37 0.45 0.33 0.42
aClean wool growth, calculated from greasy wool growth multiplied by yield taken from Table 3. Greasy
wool growth data are dyeband results for weeks 5±8.
bCalculated as 0.16protein accretion according to AFRC (1993) (Equation (94)). cN concentration in clean wool taken from Table 4 for weeks 5±8.
equation (94) from AFRC, 1993), and N retention in clean wool was calculated based on measured N concentration and total CWG. Efficiency of utilisation of MP for N retention ranged from 0.33 to 0.45 depending on the treatments and was in between the efficiency assumed for body protein and wool retention. Assuming a level of protection of 65% for methionine from Lactet (Muramatsu et al., 1994), SAA supply with MP was approximately 1.5 and 3.5 g per day for unsupplemented and methionine supplemented diets. With supplementation of rumen protected methionine, the amino acid pattern in microbial protein obviously improved in relation to the requirement as the efficiency of utilisation of MP increased by 13, 22 and 27% for diets containing untreated, heat- and formaldehyde-treated lupins (Table 8). This demonstrates the role that the amino acid pattern of MP plays in the utilisation of MP. In terms of improvement of performance prediction, protein evaluation systems need to be further developed in order to consider changes in amino acid pattern of MP due to changing proportions of undegraded protein, due to differences in feed amino acid pattern and due to the supplementation of individual rumen protected amino acids, particularly when designed for production systems where one amino acid becomes so clearly limiting as methionine for wool production.
5. Conclusion
Treating lupins with formaldehyde or heat cannot be recommended as means to improve the lupin protein quality for wool production unless extra rumen protected methionine is supplemented.
Acknowledgements
This study was funded in part by the Australian woolgrowers through the CRC for Premium Wool Quality. Markus Rodehutscord was fellow of the Deutsche Forschungs-gemeinschaft (DFG). We thank Dres. John Ashes and Suresh Gulati, CSIRO Div. of Animal Production Prospect, for treating the lupins with formaldehyde and Degussa AG, Hanau, for amino acid analysis.
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