Effects of abomasal infusion of cottonseed oil and dietary enzyme supplementation on dairy goats

Three lactating multiparous Sannen dairy goats in late lactation (146±7 days of milk) were used in a 3×3 Latin square design to determine the effects of abomasal infusion of cottonseed oil and dietary enzyme supplementation on milk yield and composition. Treatments were twice daily abomasal infusion of 1. 50 ml d-1 water plus 5 g kemzyme enzyme kg-1 dry matter (DM) of feed (ENZ), 2. 50 g cottonseed oil d-1 (OIL), and 3. 50 g cottonseed oil d-1 plus 5 g kemzyme enzyme kg-1 DM of feed (ENZ+OIL). Goats were fed a basal diet of 40% lucerne hay and 60% concentrates offered ad libitum. Each experimental period consisted of 14 days of adaptation and 5 days of total collection. Abomasal infusion of cottonseed oil decreased (P<0.05) DM intake and increased (P<0.05) milk fat percentage. Dietary supplementation of enzyme in the diet increased (P<0.05) DM and organic matter (OM) intake but had no effect (P>0.05) on apparent digestibility of DM, OM, crude protein, acid detergent fibre, neutral detergent fibre and ether extract. Cholesterol and plasma triglycerides increased (P<0.05) with cottonseed oil infusion but infusion of cottonseed oil with dietary enzyme supplementation had no (P>0.05) added benefit on lactational performance of the goats. These results suggest that abomasal infusion of cottonseed oil increased the amount of lipids reaching the mammary tissues and therefore increased milk fat percentage without altering milk yields.


INTRODUCTION
Several studies have shown that fibrolytic enzymes can increase milk yield in dairy cows fed forage-based (Schingoethe et al., 1999;Yang et al., 1999), and concentrate-based diets (Rode et al., 1999).Several studies have reported no effects of fibrolytic enzymes on milk yield (e.g., Rode et al., 1999;Kung et al., 2002) whereas other have reported negative effects (Dhiman et al., 2002;Knowlton et al., 2002).Increased dry matter intake (DMI) with dietary enzyme supplementation have been observed in several studies (e.g., Lewis et al., 1999;Beauchemin et al., 2000).However, other authors have also reported no effects of dietary enzyme supplementation on DMI (Rode et al., 1999;Kung et al., 2000Kung et al., , 2002)).Although the reasons for these conflicting results are unknown, it has been suggested that the increased DMI observed in some studies with dietary enzyme supplementation may have been due to increased palatability or higher rate of passage of feed through the digestive tract (Beauchemin et al., 2000).
In lactating dairy cows, it was shown that when 1.0 to 1.1 kg/d of rape seed oil was infused directly into the proximal duodenum, oil-free DM intake was lower in oil-infused than in control cows (Gagliostro and Chilliard, 1991).It has also been shown that abomasal infusion of unsaturated long-chain fatty acid decreased DMI (Drackley et al., 1992;Benson et al., 2001;Litherland et al., 2005).However, little work has been done to investigate the combined effect of oil infusion and dietary enzyme supplementation in ruminants.The objective of this study, therefore, was to determine the effects of abomasal infusion of cottonseed oil and dietary enzyme supplementation on DMI, plasma metabolite and milk yield and composition in dairy goats.

Animals and treatment
The experiment was conducted at the Animal Research Station, Ferdowsi University of Mashhad (Iran).Three multiparous Saanen dairy goats in late lactation (146±7 days in milk; 40±3.8 kg of body weight) were fitted with flexible abomasal cannulae and placed in individual crates in a 3×3 Latin square design.The treatments consisted of twice daily abomasal infusion of 1. 50 ml d -1 water plus 5 g kemzyme enzyme kg -1 DM of feed (ENZ), 2. 50 g cottonseed oil d -1 (OIL), and 3. 50 g cottonseed oil d -1 plus 5 g kemzyme enzyme kg -1 DM of feed (ENZ+OIL).Each experimental period consisted of 19 days; 14 days of dietary adaptation and 5 days of milk, rumen fluid and blood sample collection.
The cottonseed oil was infused in two equal proportions at each feeding whereas the enzyme was top-dressed on the total mixed ration (TMR) 24 h before feeding.The enzyme used was a powdered multi-enzyme feed additive called Kemzyme (Kemin, Belgium) obtained commercially.According to the manufacturer, the supplement contained 5000 units (U) of cellulase, 20000 U xylanase, 3000 U ß-glucanase, 450 U protease, 540 U amylase and 100 U lipase per gram of enzyme.The goats were fed a basal diet of 40% lucerne hay and 60% concentrates offered as a TMR for ad libitum intake in two equal meals at 08.00 and 20.00 h.The ingredient and chemical composition of the basal diet is presented in Table 1.Clean, fresh water was available at all time.Before the start of the experiment, the goats were treated against internal parasites and vaccinated for enterotoxaemia.

Sampling and analysis
At 08.00 h of each day after the collection of orts, the goats were hand-milked and individual milk yields recorded and the goats offered their feed allocation for the day.A sample of the milk was also obtained for compositional analysis (Milkoscan 605, Foss, Denmark).The feed offered and orts for each goat were recorded and a sub-sample of approximately 10% was collected and stored at -20ºC until the end of the experiment for subsequent analysis.On d 5 of the collection period, blood was obtained by venipuncture of the jugular vein into EDTA-containing vacuum tubes (Vacutainer, Becton-Dickinson, Rutherford, NJ) 2 h after feeding.The blood samples were immediately cooled in ice and then centrifuged at 3000 g for 15 min to obtain plasma which was stored at -20°C until analysis.The plasma samples were analysed for glucose and urea nitrogen using the enzymatic method of Sigma Diagnostics (St. Louis, MO) and triglycerides and cholesterol by infrared spectrophotometry.Rumen fluid samples were also collected from each goat on d 5 using a Geishauser oral probe (Geishauser, 1993) and strained through two layer of cheesecloth.The pH of the rumen fluid was then determined using a pH meter (Metrohm, 691, Swiss).Rumen liquor was centrifuged at 3000 g for 10 min and a sub-sample of 10 ml analysed for NH 3 -N (Kjeltec, 2300, Auto Analyzer, Tecator, Sweden).
During the collection period, total faeces was collected each day and weighed.The faeces was thoroughly mixed and a sub-sample taken and stored at -20 o C for later analysis.Urine was collected in plastic bottles under acid conditions by adding concentrated sulphuric acid (Fisher Scientific, Fairlawn, NJ) to the empty polyethylene urine collection bottles daily.The urine was mixed and aliquots (10%) were taken each day for analysis.The feed and faecal samples were ovendried at 60°C to constant weight and then ground to pass through a 1-mm screen and analysed for dry matter (DM), organic matter (OM), and ether extract (EE) according to AOAC (1990) and acid detergent fibre (ADF) and neutral detergent fibre (NDF) according to Van Soest et al. (1991).Non-casein N was determined by Kjeldahl analysis of the filtrate after precipitation with 10% acetic acid and 1N sodium acetate and subsequent determination of the N content.Casein N was calculated as the difference between total N and non casein N. True protein was calculated as the difference between total N and NPN, and whey protein calculated as the difference between true protein and casein (Rawland, 1938).The apparent digestibility coefficients of DM, OM, CP, EE, NDF, and ADF were calculated as ((intake-output)/intake).Nitrogen balance was calculated as described by Rawland (1938).

Statistical analysis
Data were analysed in a 3×3 Latin square design using the general linear model procedures of SAS (SAS, 1999).The model was: where: Y ijk = observation, µ = overall mean, G i = goat effect, P j = period effect, T k = treatment effect, E ijk = random error.Effects were considered significant at P<0.05.

RESULTS AND DISCUSSION
Nutrient intake and apparent digestibility coefficients are presented in Table 2. Enzyme supplementation increased (P<0.05) total DMI and OM intake compared to oil infusion (OIL) (Table 2) in agreement with previous studies (e.g., Lewis et al., 1999;Beauchemin et al., 2000).However, other studies did not observe any enzyme effect on DMI (Rode et al., 1999;Kung et al., 2000Kung et al., , 2002)).These contrasting results in response to enzyme supplementation are probably  and b) in a row differ significantly (P<0.05) according to Tukey test; SEM -standard error of the mean due to differences in diet composition (Beauchemin et al., 1999) and enzyme application methods (Yang et al., 2000).The (ENZ+OIL) treatment was intermediate between enzyme supplementation and oil infusion but was not different (P>0.05) from the other two treatment groups.Although the mechanism with which the abomasal oil infusion reduces DMI is not fully understood, it has been suggested that this reduction in intake was probably due to increased level of some circulating gut peptide such as cholecystokinin (Palmquist, 1994;Benson et al., 2001) or increased plasma concentrations of glucagon-like peptide 1 (Litherland et al., 2005), neither of which were measured in this experiment.
There were no treatment differences (P>0.05) in DM, OM, CP, NDF, ADF, and EE digestibility (Table 2).Although most of the published literature have reported increased total tract digestibility of DM, OM or both following treatment with a fibrolytic enzyme mixture (e.g., Rode et al., 1999;Yang et al., 1999;Beauchemin et al., 2000), other studies have reported no effect of exogenous fibrolytic enzyme on digestibility DM or OM (e.g., Lewis et al., 1999;McAllister et al., 1999McAllister et al., , 2000;;Yang et al., 2000) in agreement with the current study.In the present study, the elevated DM and OM intakes and unchanged apparent digestibility coefficient of the diet (Table 2) was probably accompanied by faster rate of ruminal and total tract rate of passage.
Milk yield and composition are presented in Table 3.There were no treatment difference (P>0.05) in milk yield and composition, except milk fat percentage.In dairy cows, Benson et al. (2001) reported that milk fat percentage was increased in response to oil infusion in midlactation, but not in early lactation and stated that when cows were in positive energy balance (such as in mid-or late lactation), supplemental fat were secreted in milk or used for body adipose tissue.There were no treatment difference (P>0.05) in fat corrected milk (FCM), protein and solids-not-fat (SNF), and efficiency (FCM/DMI; Table 3).Total solids, lactose, casein, true protein, NPN, whey protein and milk urea N were also not affected (P>0.05) by treatments in agreement with previous studies (e.g., Stokes, 1992) that have reported no change in milk composition when cows were fed enzyme-treated forages.The observation that the estimated daily NE L intake was unchanged by oil infusion in the present study suggests that DMI was being metabolically regulated to achieve energy homeostasis.
Rumen fermentation characteristic was relatively stable as evidenced by the lack of treatment effect (P>0.05) on rumen pH, ammonia nitrogen, and fibre digestibility in agreement with previous studies.There were no treatments effects (P>0.05) on blood glucose, albumin and BUN concentrations.It has been reported that oil infusion in dairy cows did not alter ruminal pH and ammonia nitrogen, or blood glucose concentration (Drackley et al., 1992).Dietary supplementations with enzymes in dairy cows have also been shown not to effect ruminal pH or ammonia nitrogen (Yang et al., 1999).Goats had significantly greater concentrations of total cholesterol (2.62, 3.07 and 2.93±0.026mmol l -1 ; mean ± SE, respectively for ENZ, OIL and ENZ+OIL) and triglyceride (0.31, 0.36 and 0.33±0.006mmol l -1 ; mean ± SE, respectively, for ENZ, OIL and ENZ+OIL) in plasma when infused with cottonseed oil.Post-ruminal infusions of rape seed oil (Gagliostro et al., 1991) or saturated or unsaturated fatty acid (Drackley et al., 1992) increased concentrations of cholesterol in the plasma of dairy cows.The intake of N, excretion of N in faeces and urine, absorbed and retained N was not affected (P>0.05) by treatment in agreement with the observation of Knowlton et al. (2002).

CONCLUSIONS
Our results show that infusion of cottonseed oil into the abomasum of lactating dairy goats decreases dry matter intake (DMI) whereas enzyme supplementation increased DMI.Abomasal infusion of cottonseed oil or supplementation of enzyme had no effect on milk yield and milk composition, despite the increased milk fat percentage.Infusions of cottonseed oil did not alter NE L intake.These results suggest that abomasal infusion of cottonseed oil increased amount of lipids reaching the mammary tissues and therefore increased milk fat percentage without altering milk yield.

Table 2 .
Nutrient intake, apparent digestibility coefficient of goats in response to dietary enzyme supplementation (ENZ), abomasal infusion of cottonseed oil (oil) or both(ENZ+OIL)

Table 3 .
Milk yield, composition and efficiency of production of goat milk in response to dietary enzyme supplementation (ENZ), abomasal infusion of cottonseed oil (OIL) or both(ENZ+OIL)