- Email: [email protected]
Effects of Abomasal Infusions of Glucose and Proprionate on Milk Yield and Composition
Effects of Abomasal Infusions of Glucose and Propionate on Milk Yield and Composition R. A. FROBISH and C. L. DAVIS Department of Dairy Science University of Illinois Urbane 61801
ABSTRACT
According to the glucogenic theory increased glucose availability is responsible for the low-milk fat problem that occurs when cows are fed high-grain, low-fiber diets. Abomasal infusions of glucose or propionate were used to test this theory. In the first trial, an average of 2.15 kg of glucose (per cow) was infused daily for 5 days. Compared to the control treatment (water infusions), the glucose infusions increased glucose concentrations of plasma (65.93 to 70.27 mg/100 ml) but had no significant effects on insulin in plasma. Milk yield increased 1.9 kg per day while the percentages of milk fat and protein decreased slightly (.31 and .14) as a result of glucose infusions. However, daily yields of milk fat and protein were not affected significantly by the increased availability of glucose. In a second trial, propionate (15 moles/cow/ day) was infused into the abomasum continuously for 5 days. Feed intake was reduced 2.53 kg per day during propiohate infusions. Milk yield, milk composition, and glucose and insulin concentrations in plasma were not affected by postruminal infusions of propionate.
INTRODUCTION
In recent years dairymen have increased grain feeding, increased the use of silage, and decreased the use of long hay as means of providing nutrients to the cow at the least cost. Often the feeding of a high ratio of readily digestible carbohydrates to fibrous constituents has changed rumen fermentation (increased propionate production) and decreased milk fat percentage (40 to 50%). Many investigators
Received August 19, 1976.
have searched for the link between alterations in the proportion of volatile fatty acids formed by rumen fermentation and milk fat percentage. Investigations into the low-milk fat problem have centered primarily around three theories. Balch et al. (2) suggested that depression in milk fat is caused by a deficiency in the amount of acetate supplied by rumen microorganisms. Van Soest and Allen (31) proposed that an increased microbial production of propionate decreases production and availability of ketone bodies, e s p e c i a l l y beta-hydroxybutyrate (BHBA), for milk fat synthesis. McClymont and Vallance (19) postulated that the glucogenic response during high propionate production increases release of insulin which suppresses mobilization of fat from tissues thereby causing a decline in blood lipids required for milk fat synthesis. Reports from this laboratory (7, 10, 23) have refuted the acetate and BHBA theories. Various researchers have attempted to test the glucogenic theory by measuring the effects of intraruminal (3, 25, 26), intravenous (14, 24, 29), or abomasal (28, 32) infusions of glucose or propionate on milk fat production. Results from these experiments have been inconclusive in supporting or refuting the glucogenic theory. Therefore, the present investigation was to test the glucogenic theory by measuring the effects of abomasally infused glucose or glucogenic compounds on milk production and composition in lactating dairy cows. EXPERIMENTAL PROCEDURE Glucose Infusion Trial
Eight rumen-fistulated cows in midlactation producing 20 to 30 kg of milk daily were used to measure the effects of abomasal infusions of glucose on milk yield, milk composition, glucose and insulin of plasma. Cows were fed 1 kg of 15%-crude protein concentrate mixture 204
MILK FAT DEPRESSION (ground shelled corn 84.25%, soybean meal 13%, dicalcium phosphate 1.50%, trace mineralized salt 1.20%, vitamins A and D .05%) for each 2.5 kg milk produced. A sufficient amount of alfalfa-grass hay was allotted to bring the ratio of concentrate to roughage of the diet to 60:40 (dry matter). Milk production for the week prior to the start of the trial guided the daily hay and concentrate allotments fed throughout the trial. Feed refusals were determined daily. A three-period, switchback design was used to compare the effects of continuous abomasal infusions of glucose or water (control). Cows were assigned randomly to treatments, and all cows received both treatments. Each test period lasted 5 days and was preceded by a 2-day adjustment. Glucose solutions and water were infused according to method of Derrig et al. (12) at 6 liters per 24 h during the test periods. The actual quantity of glucose infused was equivalent to 1.5 times the daily lactose yield by the cow and ranged from 1.6 to 2.8 kg daily (av = 2.15 kg). Average daily milk yield for the week preceding the start of the trial was used to determine the amount of glucose infused. Infusate solutions were prepared daily and maintained at 4 C during the infusion process. Water was infused at a reduced rate during the adjustment periods to prevent blockage of the infusion tube by abomasal contents. Significant differences between treatments were determined by analysis of variance for a switchback design (9). Cows were milked twice daily, and a daily composite milk sample was obtained from each cow. Percentages of milk fat and protein were determined by the Babcock procedure and Kjeldahl method. Blood samples were collected approximately 1 h post-feeding from the tail vein on days 1 to 3 and from the internal iliac artery on days 4 and 5 of each period. All blood samples were collected in heparinized tubes, centrifuged, and the plasma was stored at 0 C until analyzed. Concentrations of plasma glucose were determined for all samples by the glucose oxidase method (Glucostat, Worthington Biochemical Corporation, Freehold, New Jersey). Plasma insulin concentrations were determined for samples collected on days 4 and 5 by a commercially prepared radioimmunoassay kit (Schwartz/Mann, Division of Becton, Dickinson and Company, Orangeburg, New
205
York). The procedure was modified slightly to give the best estimation of plasma insulin. Total volume o f the assay was increased to 400/~1, so that 200 /.d of plasma could be included, and time allowed for the binding reactions was increased to 24 h. Rumen fluid samples were collected daily (2 h post feeding) through the rumen cannula; they were acidified and stored at 0 C until analyzed. Volatile fatty acids were determined according to the procedure of Erwin et al. (13) with an Aerograph, Model 600, gas chromatograph equipped with a hydrogen flame ionization detector. Propionate Infusion Trial
Four rumen fistulated cows in midlactation, producing 25 to 35 kg of milk daily, were used to measure the effects of abomasal infusions of propionate on milk yield, milk composition, glucose and insulin of plasma. Feeding regimen, experimental design, infusion technique, and methods of measuring the aforementioned parameters were-the same for this trial as for the glucose infusion trial. In the present trial, a continuous abomasal infusion of 15 moles of porpionate was compared with a water infusion (control). A 50% sodium hydroxide solution (250 ml) was mixed with the propionic acid to raise the pH of the infusate to 5.0 to 5.2, and water was added to give a final volume of 5.1. Both infusates were administered at 5 liters per 24 h. Fecal samples were collected from all cows on the last day of the trial. Samples were extracted with water (1:1, vol:wt), the filtrate acidified, and analyzed for propionate concentrations by gas chromatography.
RESULTS A N D DISCUSSION Glucose Infusion Trial
In studies of this kind it is important to assure that the infusion does not alter the basal conditions o f feed intake and the proportions of the ruminal fermentation acids. Data in Table 1 show that feed intake was unaltered by abomasal infusion of glucose and that the rumen fermentation pattern was typical of that in cows producing milk of normal fat content as judged by a rumen acetate to propionate ratio of approximately 3 to 1. Therefore, any Journal of Dairy Science Vol. 60, No. 2
206
FROBISH AND DAVIS
TABLE 1. Effects of abomasal infusions of glucose on feed intake and rumen acetate to propionate ratio. Treatmen~
Concentrate intake (kg/day) Hay intake (kg/day) Total feed intake (kg/day) Rumen acetate/propionate ratio (molar %)
Control
Glucose
11.59± .71 a 7.31± .41 18.90 ± 1.11 3.29± .19
11.61±.58 6.78±.33 18.39 ± .74 2.97±.20
avalues represent the means -+SE for the periods during which glucose or water (control) were infused.
significant treatment difference can be attributed direcdy to an increase in glucose availability. Abomasal infusions of glucose decreased milk fat percentage (P<.05) (Table 2); however, the decrease was small (9.4%) compared to the decrease (40 to 60%) when high-grain, low-fiber diets are fed. More importandy, milk fat yield was not altered significantly by the glucose infusions showing that the decrease in milk fat percent was the result of a significant increase in milk yield. This pattern of response is n o t typical of the classical low-milk fat syndrome where there is both a decrease in percent and yield of fat. Aside from the main issue, milk protein percent was decreased, but total milk protein yield was unaffected by the infusion of glucose. Abomasal infusion of glucose increased (P<.05) the concentration of glucose in plasma, a finding which was not totally unexpected. The plasma values for glucose are within normal
physiological limits and fairly typical of those reported for cows fed high-grain, low-fiber diets (1, 11). Reports linking glucose availability and reduced milk fat percent involved intravenous infusions of glucose resulting in abnormally high blood glucose (19, 24). Plasma insulin, although not statistically different, tended to be higher for the cows receiving glucose infusions. Insulin values are lower than values reported for cattle (30); however, the comparison between treatments was valid. McAtee and Trenkle (18) observed a marked stimulation in insulin release when glucose was infused intravenously over 2 h. The route of administration and length of infusion may be responsible for the difference in results. The low correlation between glucose concentration in blood and plasma insulin in ruminants (15, 16, 18) and the absence of a substantial decrease in triglyceride of serum in cows fed a high-grain, low-fiber diet (17, 27, 33) creates doubts about an insulin involvement in the
TABLE 2. Effects of abomasal infusions of glucose on milk yield, milk composition, and plasma glucose and insulin concentrations. Treatments Parameter measured Daily milk production (kg) Milk fat percentage (%) Daily milk fat yield (g) Milk protein percentage (%) Daily milk protein yield (g) Plasma glucose (rag %) Plasma insulin (#U/ml)
Control 26.73 ± 1.76a** 3.29 +- .09* 865 ± 40 3.50 ± .06* 932 ± 59 65.93 ± .67* 9.46 ± .82
Glucose 28.63± 1.79"* 2.98 -+ .06" 843 ± 46 3.36 ± .09* 954 ± 56 70.27± 1.22" 13.36 ± 2.05
aValues represent the means ± SE for the periods during which glucose or water (control) were infused. *Significant at .05 level. **Significant at .01 level. Journal of Dairy Science Vol. 60, No. 2
MILK FAT DEPRESSION low-milk fat syndrome. Massive injections of insulin into cows receiving a normal diet failed to depress milk fat percent (5). On the other hand, changes in activities of specific enzyme systems associated with milk fat synthesis (4, 8, 21) appear to support the idea of an insulin involvement although how is not clear. Questions may be raised logically as to 1) how much of the infused glucose was absorbed; and 2) did the infusion of glucose abomasally increase the availability of glucose during the infusion of glucose. 0rskov et al. (22) reported that sheep feces became increasingly fluid when glucose was supplied in excess of the absorptive capacity of the gut. In no instance did the feces excreted by the cows change in switching from the control (water) infusion to glucose infusion. Bartley and Black (6) demonstrated that the administration of exogenous glucose depressed endogenous glucose production (gluconeogenesis) resulting in only a slight increase in glucose availability to the animal. It is possible that gluconeogenesis was reduced in the present experiment when glucose was supplied abomasaUy; however, plasma glucose was increased significantly by the exogenous supply of glucose, and that in plasma was comparable to amounts reported for cows fed a high-grain, low-fiber diet. Furthermore, the fact that the cows receiving glucose infusions increased their milk yield argues for greater amounts o f glucose available to them. Propionate Infusion Trial
The observed depression in milk fat yield when cows are fed high-grain, low-fiber diet has been more highly correlated with the proportion of rumen propionate than acetate (20).
207
Rumen propionate production was increased two-fold when cows were fed a high-grain, low-fiber diet as compared to a normal diet (7). Moreover, in cattle (18) intravenous infusions of propionate stimulated greater insulin release than intravenous infusions of glucose. These findings raised the possibility that increased propionate availability, not glucose availability, is the cause of the low-milk fat problem when cows are fed high-grain, low-fiber diets. Another experiment was initiated to study the effects of abomasally infused propionate on milk fat yield and composition. Total feed intake decreased (P<.05) during propionate infusions (Table 3). Majority of this decrease resulted from an average reduction of 1.83 kg/cow/day (P<.05) in concentrate consumption. There was no significant difference in hay consumption between treatments. Other workers have found that feed intake was not affected by intravenous administrations of propionate (14, 29); however, feed refusals have been reported (26) during intraruminal infusion of large amounts of propionate (greater than 1600 ml per day). The slight reduction in feed intake in our study did not alter rumen fermentation. Ratios of acetate to propionate in the rumen were normal throughout the trial (Table 3). Therefore, any significant difference in the parameters measured could be attributed to increased propionate availability. Postruminal administration of propionate (15 moles) had no significant effect on milk yield or composition as compared to the control treatment (Table 4). In earlier studies (3, 25, 26) intraruminal infusions of propionate also failed to increase milk production significantly or to decrease milk fat yield. In contrast, Fisher and Elliot (14) reported an increase (P<.05) in milk production and a decrease
TABLE 3. Effects of abomasal infusions of propionate on feed intake and rumen acetate to propionate ratio.
Concentrate intake (kg/day) Hay intake (kg/day) Total feed intake (kg/day) Rumen acetate/propionate ratio (molar %)
Control
Propionate
11.65-+.57 a* 8.25 + .35 19.90 +- .92* 3.66 ± .22
9.82± .79* 7.55 +- .50 17.37 ± 1.27" 3.77 ± .14
avalues represent the means -+SE for the periods during which propionate or water (control) were infused. *Significant at .05 level. Journal of Dairy Science Vol. 60, No. 2
208
FROBISH AND DAVIS
TABLE 4. Effects of abomasal infusions of propionate on milk yield, milk composition, and plasma glucose and insulin concentrations. Parameter measured
Control
Propionate
Daily milk production (kg) Milk fat percentage (%) Daily milk fat yield (g) Milk protein percentage (%) Daily milk protein yield (g) Plasma glucose (mg %) Plasma insulin (gU/ml)
23.88 +- 1.97 a 3.32 -+ .07 789 ,+ 58 3.42 +- .05 813 +- 61 71.70,+ 3.66 12.17,+ 1.25
24.25 ,+ 1.86 3.19 +- .07 772 ± 54 3.35 -+ .09 807 ,+ 49 67.53-+ 1.45 10.89+- 1.97
avalues represent the means +- SE for the periods during which propionate or water (control) were infused.
(P<.05) in milk fat percentage w h e n smaJler quantities of p r o p i o n a t e were infused intravenously. The lack o f an increase in milk protein yield during periods of p r o p i o n a t e infusions does n o t agree with the consistent increases observed by o t h e r researchers (14, 25, 26). Glucose and insulin c o n c e n t r a t i o n s in plasma were u n c h a n g e d by the infusion of p r o p i o n a t e (Table 4). T h e glucose data agree with earlier observations o f Fisher and Elliot (14). The lack of increase in plasma insulin during periods of p r o p i o n a t e infusions does n o t agree with the report by M c A t e e and Trenkle (18) that propionate infusions increased plasma insulin. However, t h e y n o t i c e d that insulin in plasma reached a m a x i m u m a b o u t 30 rain after initiation of the infusion and then declined, even though the infusion was continued. In our study, plasma insulin may have increased shortly after initiation o f the infusion but declined to preinfusion c o n c e n t r a t i o n s by the time the b l o o d samples were collected. Cows receiving a normal diet ( 6 0 : 4 0 concentrate to roughage ratio) were supplied postruminally with an additional 15 moles of p r o p i o n a t e per day. It can be assumed that the additional propionate was absorbed since fecal samples f r o m cows receiving p r o p i o n a t e infusions did n o t c o n t a i n an increased a m o u n t of p r o p i o n a t e as c o m p a r e d to fecal samples f r o m cows receiving water infusions. Postruminal addition of 15 moles of p r o p i o n a t e to the 13 moles n o r m a l l y p r o d u c e d in the r u m e n o f cows fed a similar diet (7) increased the total available p r o p i o n a t e to levels similar to p r o d u c t i o n o f p r o p i o n a t e in the r u m e n w h e n cows are fed a high-grain, lowfiber diet. However, milk yield and c o m p o s i t i o n were unchanged by the abomasal infusions of Journal of Dairy Science Vol. 60, No. 2
propionate. Therefore, under normal r u m e n ferm e n t a t i o n , increased p r o p i o n a t e availability, per se, is n o t the cause of the low milk fat syndrome. REFERENCES
1 Annison, E. F., R. Bickerstaffe, and J. L. Linzell. 1974. Glucose and fatty acid metabolism in cows producing milk of low fat content. J. Agr. Sci., Camb. 82:87. 2 Balch, C. C., D. A. Balch, S. Bartlett, M. P. Bartrum, V. W. Johnson, S. J. Rowland, and J. Turner. 1955. Studies of the secretion of milk of low fat content by cows on diets low in hay and high in concentrates. VL The effect on the physical and biochemical processes of the reticulo-rumen. J. Dairy Res. 22:270. 3 Balch, C. C., W. H. Broster, V. W. Johnson, C. Line, J. A. F. Rook, J. D. Sutton, and V. J. Tuck. 1967. The effect of milk yidd and composition of adding the calcium salts of acetic, propionic, butyric, and lactic acids to the diets of dairy cows. J. Dairy Res. 34:199. 4 Baldwin, R. L., H. J. Lin, W. Cheng, R. Cabrera, and M. Ronning. 1969. Enzyme and metabolite levels in mammary and abdominal adipose tissue of lactating dairy cows. J. Dairy Sci. 52:183. 5 Baldwin, R. L., J. R. Reichl, S. Louis, N. E. Smith, g. T. Yang, and E. Osborne. 1973. Effects of age, pregnancy, and lactation on rat, guinea pig, and cow adipose enzyme activities and cow adipose metabolism. J. Dairy Sci. 56: 340. 6 Bartley, J. C., and A. L. Black. 1966. Effect of exogenous glucose on glucose metabolism in dairy cows. J. Nutr. 89:317. 7 Bauman, D. E., C. L. Davis, and H. F. Bucholtz. 1971. Propionate production in the rumen of cows fed either a control or high-grain, low-fiber diet. J. Dairy Sci. 54:1282. 8 Benson, J. E., E. W. Askew, R. S. Emery, and J. W. Thomas. 1972. Metabolism of fatty acids by adipose tissue and liver of cows fed normal, restricted roughage or MgO supplemented rations. J. Dairy Sci. 55:83.
MILK FAT DEPRESSION 9 Brandt, A. E. 1938. Tests of significance in reversal of switchback trials. Iowa St. At. Exp. Sta. Bull. 234:58. 10 Davis, C. L. 1967. Acetate production in the rumen fo cows fed either control or low-fiber, high-grain diets. J. Dairy Sci. 50:1621. i1 Davis, C. L., and R. E. Brown. 1970. Low-fat milk syndrome. Physiology of Digestion and Metabolism in the Ruminant. ed. A. T. Phillipson, Griel Press, New Castle. 12 Derrig, R. G., J. H. Clark, and C. L. Davis. 1974. Effect of abomasal infusion of sodium caseinate on milk yield, nitrogen utilization and amino acid nutrition of the dairy cows. J. Nutr. 104:151. 13 Erwin, E. S., G. J. Marco, and E. M. Emery. 1961. Volatile fatty acid analyses of blood and rumen fluid by gas chromatography. J. Dairy Sci. 44:1768. 14 Fisher, L. J., and J. M. Elliot. 1966. Effect of intravenous infusion of propionate or glucose on bovine milk composition. J. Dairy Sci. 49:826. 15 Horino, M., L. J. Machlin, F. Hertelendy, and D. M. Kipnis. 1968. Effect of short-chain fatty acids on plasma insulin in ruminant and nonruminant species. Endocrinology 83:118. 16 Hove, K., and A. K. Biota. 1973. Plasma insulin and growth hormone in dairy cows; diurnal variation and relation to food intake and plasma sugar and acetoacetate levels. Acta Endocrinol. 73:289. 17 Jenny, B. F., C. E. Polan, and P. T. Chandler. 1972. Acetate and stearate utilization by lactating cows fed high grain-restricted roughage rations. J. Dairy Sci. 55:1481. 18 McAtee, J. W., and A. Trenlde. 1971. Metabolic regulation of plasma insulin levels in cattle. J. Anim. Sci. 33:438. 19 McClymont, G. L., and S. Vallance. 1962. Depression of blood glycerides and milk fat synthesis by glucose infusion. Proc. Nutr. Sci. 21 :xli. 20 McCullough, M. E. 1966. Relationships between rumen fluid volatile fatty acids and milk fat percentage and feed intake. J. Dairy Sci. 49:896. 21 Opstvedt, J., R. L. Baldwin, and M. Ronning. 1967. Effect of diet upon activities of several enzymes in abdominal adipose and mammary tissues in the lactating dairy cow. J. Dairy Sci. 50:108.
209
22 Orskov, E. R., R. W. Mayes, and A. Penn. 1971. The capacity for the removal of gtucose from the small intestine by mature sheep. Proc. Nutr. Soc., 30:43a. 23 Palmquist, D. L., C. L. Davis, R. E. Brown, and D. S. Sachan. 1969. Availability and metabolism of various substrates in ruminants. V. Entry rate into the body and incorporation into milk fat of D(-)beta-hydroxybutyrate. J. Dairy Sci. 52:633. 24 Rao, D. R., G. E. Hawkins, and R. C. Smith. 1973. Effect of glucose and insulin on lipoprotein lipase activity in adipose tissue and milk. J. Dairy Sci. 56:1415. 25 Rook, J. A. F., and C. C. Balch. 1961. The effects of intraruminal infusions of acetic, propionic and butyric acids on the yield and composition of the milk of the cows. Br. J. Nutr. 15:361. 26 Rook, J. A. F., C. C. Balch, and V. W. Johnson. 1965. Further observations on the effects of intraruminal infusions of volatile fatty acids and of lactic acid on the yield and composition of the milk of the cow. Br. J. Nutr. 19:93. 27 Satter, L. D., and A. N. Bringe. 1969. Effect of abrupt ration changes on milk and blood components. J. Dairy Sci. 52:1776. 28 Spires, H. R. 1974. Effect of postruminal infusions of glucose and sodium caseinate on milk production, amino acid utilization and glucose synthesis of lactating cows. M. S. Thesis. University of Illinois, Urbana. 29 Storry, J. E., and J. A. F. Rook. 1965. Effects of intravenous infusions of acetate, beta-hydroxybutyrate, triglyceride and other metabolites on the composition of the milk fat and blood in cows. Biochem. J. 97:879. 30 Trenkle, A. 1972. Radioimmunoassay of plasma hormones: Review of plasma insulin in ruminants. J. Dairy Sci. 55:1200. 31 Van Soest, P. J., and N. N. Allen. 1959. Studies on the relationships between rumen acids and fat metabolism of ruminants fed on restricted roughage diets. J. Dairy Sci. 42:1977. 32 Vik-Mo, L., R. S. Emery, and J. T. Huber. 1974. Milk protein production in cows abomasally infused with casein or glucose. J. Dairy Sci. 57:869. 33 Yang, Y. T., and R. L. Baldwin. 1973. Lipolysis in isolated cow adipose cells. J. Dairy Sci. 56: 366.
Journal of Dairy Science Vol. 60, No. 2
ABSTRACT
According to the glucogenic theory increased glucose availability is responsible for the low-milk fat problem that occurs when cows are fed high-grain, low-fiber diets. Abomasal infusions of glucose or propionate were used to test this theory. In the first trial, an average of 2.15 kg of glucose (per cow) was infused daily for 5 days. Compared to the control treatment (water infusions), the glucose infusions increased glucose concentrations of plasma (65.93 to 70.27 mg/100 ml) but had no significant effects on insulin in plasma. Milk yield increased 1.9 kg per day while the percentages of milk fat and protein decreased slightly (.31 and .14) as a result of glucose infusions. However, daily yields of milk fat and protein were not affected significantly by the increased availability of glucose. In a second trial, propionate (15 moles/cow/ day) was infused into the abomasum continuously for 5 days. Feed intake was reduced 2.53 kg per day during propiohate infusions. Milk yield, milk composition, and glucose and insulin concentrations in plasma were not affected by postruminal infusions of propionate.
INTRODUCTION
In recent years dairymen have increased grain feeding, increased the use of silage, and decreased the use of long hay as means of providing nutrients to the cow at the least cost. Often the feeding of a high ratio of readily digestible carbohydrates to fibrous constituents has changed rumen fermentation (increased propionate production) and decreased milk fat percentage (40 to 50%). Many investigators
Received August 19, 1976.
have searched for the link between alterations in the proportion of volatile fatty acids formed by rumen fermentation and milk fat percentage. Investigations into the low-milk fat problem have centered primarily around three theories. Balch et al. (2) suggested that depression in milk fat is caused by a deficiency in the amount of acetate supplied by rumen microorganisms. Van Soest and Allen (31) proposed that an increased microbial production of propionate decreases production and availability of ketone bodies, e s p e c i a l l y beta-hydroxybutyrate (BHBA), for milk fat synthesis. McClymont and Vallance (19) postulated that the glucogenic response during high propionate production increases release of insulin which suppresses mobilization of fat from tissues thereby causing a decline in blood lipids required for milk fat synthesis. Reports from this laboratory (7, 10, 23) have refuted the acetate and BHBA theories. Various researchers have attempted to test the glucogenic theory by measuring the effects of intraruminal (3, 25, 26), intravenous (14, 24, 29), or abomasal (28, 32) infusions of glucose or propionate on milk fat production. Results from these experiments have been inconclusive in supporting or refuting the glucogenic theory. Therefore, the present investigation was to test the glucogenic theory by measuring the effects of abomasally infused glucose or glucogenic compounds on milk production and composition in lactating dairy cows. EXPERIMENTAL PROCEDURE Glucose Infusion Trial
Eight rumen-fistulated cows in midlactation producing 20 to 30 kg of milk daily were used to measure the effects of abomasal infusions of glucose on milk yield, milk composition, glucose and insulin of plasma. Cows were fed 1 kg of 15%-crude protein concentrate mixture 204
MILK FAT DEPRESSION (ground shelled corn 84.25%, soybean meal 13%, dicalcium phosphate 1.50%, trace mineralized salt 1.20%, vitamins A and D .05%) for each 2.5 kg milk produced. A sufficient amount of alfalfa-grass hay was allotted to bring the ratio of concentrate to roughage of the diet to 60:40 (dry matter). Milk production for the week prior to the start of the trial guided the daily hay and concentrate allotments fed throughout the trial. Feed refusals were determined daily. A three-period, switchback design was used to compare the effects of continuous abomasal infusions of glucose or water (control). Cows were assigned randomly to treatments, and all cows received both treatments. Each test period lasted 5 days and was preceded by a 2-day adjustment. Glucose solutions and water were infused according to method of Derrig et al. (12) at 6 liters per 24 h during the test periods. The actual quantity of glucose infused was equivalent to 1.5 times the daily lactose yield by the cow and ranged from 1.6 to 2.8 kg daily (av = 2.15 kg). Average daily milk yield for the week preceding the start of the trial was used to determine the amount of glucose infused. Infusate solutions were prepared daily and maintained at 4 C during the infusion process. Water was infused at a reduced rate during the adjustment periods to prevent blockage of the infusion tube by abomasal contents. Significant differences between treatments were determined by analysis of variance for a switchback design (9). Cows were milked twice daily, and a daily composite milk sample was obtained from each cow. Percentages of milk fat and protein were determined by the Babcock procedure and Kjeldahl method. Blood samples were collected approximately 1 h post-feeding from the tail vein on days 1 to 3 and from the internal iliac artery on days 4 and 5 of each period. All blood samples were collected in heparinized tubes, centrifuged, and the plasma was stored at 0 C until analyzed. Concentrations of plasma glucose were determined for all samples by the glucose oxidase method (Glucostat, Worthington Biochemical Corporation, Freehold, New Jersey). Plasma insulin concentrations were determined for samples collected on days 4 and 5 by a commercially prepared radioimmunoassay kit (Schwartz/Mann, Division of Becton, Dickinson and Company, Orangeburg, New
205
York). The procedure was modified slightly to give the best estimation of plasma insulin. Total volume o f the assay was increased to 400/~1, so that 200 /.d of plasma could be included, and time allowed for the binding reactions was increased to 24 h. Rumen fluid samples were collected daily (2 h post feeding) through the rumen cannula; they were acidified and stored at 0 C until analyzed. Volatile fatty acids were determined according to the procedure of Erwin et al. (13) with an Aerograph, Model 600, gas chromatograph equipped with a hydrogen flame ionization detector. Propionate Infusion Trial
Four rumen fistulated cows in midlactation, producing 25 to 35 kg of milk daily, were used to measure the effects of abomasal infusions of propionate on milk yield, milk composition, glucose and insulin of plasma. Feeding regimen, experimental design, infusion technique, and methods of measuring the aforementioned parameters were-the same for this trial as for the glucose infusion trial. In the present trial, a continuous abomasal infusion of 15 moles of porpionate was compared with a water infusion (control). A 50% sodium hydroxide solution (250 ml) was mixed with the propionic acid to raise the pH of the infusate to 5.0 to 5.2, and water was added to give a final volume of 5.1. Both infusates were administered at 5 liters per 24 h. Fecal samples were collected from all cows on the last day of the trial. Samples were extracted with water (1:1, vol:wt), the filtrate acidified, and analyzed for propionate concentrations by gas chromatography.
RESULTS A N D DISCUSSION Glucose Infusion Trial
In studies of this kind it is important to assure that the infusion does not alter the basal conditions o f feed intake and the proportions of the ruminal fermentation acids. Data in Table 1 show that feed intake was unaltered by abomasal infusion of glucose and that the rumen fermentation pattern was typical of that in cows producing milk of normal fat content as judged by a rumen acetate to propionate ratio of approximately 3 to 1. Therefore, any Journal of Dairy Science Vol. 60, No. 2
206
FROBISH AND DAVIS
TABLE 1. Effects of abomasal infusions of glucose on feed intake and rumen acetate to propionate ratio. Treatmen~
Concentrate intake (kg/day) Hay intake (kg/day) Total feed intake (kg/day) Rumen acetate/propionate ratio (molar %)
Control
Glucose
11.59± .71 a 7.31± .41 18.90 ± 1.11 3.29± .19
11.61±.58 6.78±.33 18.39 ± .74 2.97±.20
avalues represent the means -+SE for the periods during which glucose or water (control) were infused.
significant treatment difference can be attributed direcdy to an increase in glucose availability. Abomasal infusions of glucose decreased milk fat percentage (P<.05) (Table 2); however, the decrease was small (9.4%) compared to the decrease (40 to 60%) when high-grain, low-fiber diets are fed. More importandy, milk fat yield was not altered significantly by the glucose infusions showing that the decrease in milk fat percent was the result of a significant increase in milk yield. This pattern of response is n o t typical of the classical low-milk fat syndrome where there is both a decrease in percent and yield of fat. Aside from the main issue, milk protein percent was decreased, but total milk protein yield was unaffected by the infusion of glucose. Abomasal infusion of glucose increased (P<.05) the concentration of glucose in plasma, a finding which was not totally unexpected. The plasma values for glucose are within normal
physiological limits and fairly typical of those reported for cows fed high-grain, low-fiber diets (1, 11). Reports linking glucose availability and reduced milk fat percent involved intravenous infusions of glucose resulting in abnormally high blood glucose (19, 24). Plasma insulin, although not statistically different, tended to be higher for the cows receiving glucose infusions. Insulin values are lower than values reported for cattle (30); however, the comparison between treatments was valid. McAtee and Trenkle (18) observed a marked stimulation in insulin release when glucose was infused intravenously over 2 h. The route of administration and length of infusion may be responsible for the difference in results. The low correlation between glucose concentration in blood and plasma insulin in ruminants (15, 16, 18) and the absence of a substantial decrease in triglyceride of serum in cows fed a high-grain, low-fiber diet (17, 27, 33) creates doubts about an insulin involvement in the
TABLE 2. Effects of abomasal infusions of glucose on milk yield, milk composition, and plasma glucose and insulin concentrations. Treatments Parameter measured Daily milk production (kg) Milk fat percentage (%) Daily milk fat yield (g) Milk protein percentage (%) Daily milk protein yield (g) Plasma glucose (rag %) Plasma insulin (#U/ml)
Control 26.73 ± 1.76a** 3.29 +- .09* 865 ± 40 3.50 ± .06* 932 ± 59 65.93 ± .67* 9.46 ± .82
Glucose 28.63± 1.79"* 2.98 -+ .06" 843 ± 46 3.36 ± .09* 954 ± 56 70.27± 1.22" 13.36 ± 2.05
aValues represent the means ± SE for the periods during which glucose or water (control) were infused. *Significant at .05 level. **Significant at .01 level. Journal of Dairy Science Vol. 60, No. 2
MILK FAT DEPRESSION low-milk fat syndrome. Massive injections of insulin into cows receiving a normal diet failed to depress milk fat percent (5). On the other hand, changes in activities of specific enzyme systems associated with milk fat synthesis (4, 8, 21) appear to support the idea of an insulin involvement although how is not clear. Questions may be raised logically as to 1) how much of the infused glucose was absorbed; and 2) did the infusion of glucose abomasally increase the availability of glucose during the infusion of glucose. 0rskov et al. (22) reported that sheep feces became increasingly fluid when glucose was supplied in excess of the absorptive capacity of the gut. In no instance did the feces excreted by the cows change in switching from the control (water) infusion to glucose infusion. Bartley and Black (6) demonstrated that the administration of exogenous glucose depressed endogenous glucose production (gluconeogenesis) resulting in only a slight increase in glucose availability to the animal. It is possible that gluconeogenesis was reduced in the present experiment when glucose was supplied abomasaUy; however, plasma glucose was increased significantly by the exogenous supply of glucose, and that in plasma was comparable to amounts reported for cows fed a high-grain, low-fiber diet. Furthermore, the fact that the cows receiving glucose infusions increased their milk yield argues for greater amounts o f glucose available to them. Propionate Infusion Trial
The observed depression in milk fat yield when cows are fed high-grain, low-fiber diet has been more highly correlated with the proportion of rumen propionate than acetate (20).
207
Rumen propionate production was increased two-fold when cows were fed a high-grain, low-fiber diet as compared to a normal diet (7). Moreover, in cattle (18) intravenous infusions of propionate stimulated greater insulin release than intravenous infusions of glucose. These findings raised the possibility that increased propionate availability, not glucose availability, is the cause of the low-milk fat problem when cows are fed high-grain, low-fiber diets. Another experiment was initiated to study the effects of abomasally infused propionate on milk fat yield and composition. Total feed intake decreased (P<.05) during propionate infusions (Table 3). Majority of this decrease resulted from an average reduction of 1.83 kg/cow/day (P<.05) in concentrate consumption. There was no significant difference in hay consumption between treatments. Other workers have found that feed intake was not affected by intravenous administrations of propionate (14, 29); however, feed refusals have been reported (26) during intraruminal infusion of large amounts of propionate (greater than 1600 ml per day). The slight reduction in feed intake in our study did not alter rumen fermentation. Ratios of acetate to propionate in the rumen were normal throughout the trial (Table 3). Therefore, any significant difference in the parameters measured could be attributed to increased propionate availability. Postruminal administration of propionate (15 moles) had no significant effect on milk yield or composition as compared to the control treatment (Table 4). In earlier studies (3, 25, 26) intraruminal infusions of propionate also failed to increase milk production significantly or to decrease milk fat yield. In contrast, Fisher and Elliot (14) reported an increase (P<.05) in milk production and a decrease
TABLE 3. Effects of abomasal infusions of propionate on feed intake and rumen acetate to propionate ratio.
Concentrate intake (kg/day) Hay intake (kg/day) Total feed intake (kg/day) Rumen acetate/propionate ratio (molar %)
Control
Propionate
11.65-+.57 a* 8.25 + .35 19.90 +- .92* 3.66 ± .22
9.82± .79* 7.55 +- .50 17.37 ± 1.27" 3.77 ± .14
avalues represent the means -+SE for the periods during which propionate or water (control) were infused. *Significant at .05 level. Journal of Dairy Science Vol. 60, No. 2
208
FROBISH AND DAVIS
TABLE 4. Effects of abomasal infusions of propionate on milk yield, milk composition, and plasma glucose and insulin concentrations. Parameter measured
Control
Propionate
Daily milk production (kg) Milk fat percentage (%) Daily milk fat yield (g) Milk protein percentage (%) Daily milk protein yield (g) Plasma glucose (mg %) Plasma insulin (gU/ml)
23.88 +- 1.97 a 3.32 -+ .07 789 ,+ 58 3.42 +- .05 813 +- 61 71.70,+ 3.66 12.17,+ 1.25
24.25 ,+ 1.86 3.19 +- .07 772 ± 54 3.35 -+ .09 807 ,+ 49 67.53-+ 1.45 10.89+- 1.97
avalues represent the means +- SE for the periods during which propionate or water (control) were infused.
(P<.05) in milk fat percentage w h e n smaJler quantities of p r o p i o n a t e were infused intravenously. The lack o f an increase in milk protein yield during periods of p r o p i o n a t e infusions does n o t agree with the consistent increases observed by o t h e r researchers (14, 25, 26). Glucose and insulin c o n c e n t r a t i o n s in plasma were u n c h a n g e d by the infusion of p r o p i o n a t e (Table 4). T h e glucose data agree with earlier observations o f Fisher and Elliot (14). The lack of increase in plasma insulin during periods of p r o p i o n a t e infusions does n o t agree with the report by M c A t e e and Trenkle (18) that propionate infusions increased plasma insulin. However, t h e y n o t i c e d that insulin in plasma reached a m a x i m u m a b o u t 30 rain after initiation of the infusion and then declined, even though the infusion was continued. In our study, plasma insulin may have increased shortly after initiation o f the infusion but declined to preinfusion c o n c e n t r a t i o n s by the time the b l o o d samples were collected. Cows receiving a normal diet ( 6 0 : 4 0 concentrate to roughage ratio) were supplied postruminally with an additional 15 moles of p r o p i o n a t e per day. It can be assumed that the additional propionate was absorbed since fecal samples f r o m cows receiving p r o p i o n a t e infusions did n o t c o n t a i n an increased a m o u n t of p r o p i o n a t e as c o m p a r e d to fecal samples f r o m cows receiving water infusions. Postruminal addition of 15 moles of p r o p i o n a t e to the 13 moles n o r m a l l y p r o d u c e d in the r u m e n o f cows fed a similar diet (7) increased the total available p r o p i o n a t e to levels similar to p r o d u c t i o n o f p r o p i o n a t e in the r u m e n w h e n cows are fed a high-grain, lowfiber diet. However, milk yield and c o m p o s i t i o n were unchanged by the abomasal infusions of Journal of Dairy Science Vol. 60, No. 2
propionate. Therefore, under normal r u m e n ferm e n t a t i o n , increased p r o p i o n a t e availability, per se, is n o t the cause of the low milk fat syndrome. REFERENCES
1 Annison, E. F., R. Bickerstaffe, and J. L. Linzell. 1974. Glucose and fatty acid metabolism in cows producing milk of low fat content. J. Agr. Sci., Camb. 82:87. 2 Balch, C. C., D. A. Balch, S. Bartlett, M. P. Bartrum, V. W. Johnson, S. J. Rowland, and J. Turner. 1955. Studies of the secretion of milk of low fat content by cows on diets low in hay and high in concentrates. VL The effect on the physical and biochemical processes of the reticulo-rumen. J. Dairy Res. 22:270. 3 Balch, C. C., W. H. Broster, V. W. Johnson, C. Line, J. A. F. Rook, J. D. Sutton, and V. J. Tuck. 1967. The effect of milk yidd and composition of adding the calcium salts of acetic, propionic, butyric, and lactic acids to the diets of dairy cows. J. Dairy Res. 34:199. 4 Baldwin, R. L., H. J. Lin, W. Cheng, R. Cabrera, and M. Ronning. 1969. Enzyme and metabolite levels in mammary and abdominal adipose tissue of lactating dairy cows. J. Dairy Sci. 52:183. 5 Baldwin, R. L., J. R. Reichl, S. Louis, N. E. Smith, g. T. Yang, and E. Osborne. 1973. Effects of age, pregnancy, and lactation on rat, guinea pig, and cow adipose enzyme activities and cow adipose metabolism. J. Dairy Sci. 56: 340. 6 Bartley, J. C., and A. L. Black. 1966. Effect of exogenous glucose on glucose metabolism in dairy cows. J. Nutr. 89:317. 7 Bauman, D. E., C. L. Davis, and H. F. Bucholtz. 1971. Propionate production in the rumen of cows fed either a control or high-grain, low-fiber diet. J. Dairy Sci. 54:1282. 8 Benson, J. E., E. W. Askew, R. S. Emery, and J. W. Thomas. 1972. Metabolism of fatty acids by adipose tissue and liver of cows fed normal, restricted roughage or MgO supplemented rations. J. Dairy Sci. 55:83.
MILK FAT DEPRESSION 9 Brandt, A. E. 1938. Tests of significance in reversal of switchback trials. Iowa St. At. Exp. Sta. Bull. 234:58. 10 Davis, C. L. 1967. Acetate production in the rumen fo cows fed either control or low-fiber, high-grain diets. J. Dairy Sci. 50:1621. i1 Davis, C. L., and R. E. Brown. 1970. Low-fat milk syndrome. Physiology of Digestion and Metabolism in the Ruminant. ed. A. T. Phillipson, Griel Press, New Castle. 12 Derrig, R. G., J. H. Clark, and C. L. Davis. 1974. Effect of abomasal infusion of sodium caseinate on milk yield, nitrogen utilization and amino acid nutrition of the dairy cows. J. Nutr. 104:151. 13 Erwin, E. S., G. J. Marco, and E. M. Emery. 1961. Volatile fatty acid analyses of blood and rumen fluid by gas chromatography. J. Dairy Sci. 44:1768. 14 Fisher, L. J., and J. M. Elliot. 1966. Effect of intravenous infusion of propionate or glucose on bovine milk composition. J. Dairy Sci. 49:826. 15 Horino, M., L. J. Machlin, F. Hertelendy, and D. M. Kipnis. 1968. Effect of short-chain fatty acids on plasma insulin in ruminant and nonruminant species. Endocrinology 83:118. 16 Hove, K., and A. K. Biota. 1973. Plasma insulin and growth hormone in dairy cows; diurnal variation and relation to food intake and plasma sugar and acetoacetate levels. Acta Endocrinol. 73:289. 17 Jenny, B. F., C. E. Polan, and P. T. Chandler. 1972. Acetate and stearate utilization by lactating cows fed high grain-restricted roughage rations. J. Dairy Sci. 55:1481. 18 McAtee, J. W., and A. Trenlde. 1971. Metabolic regulation of plasma insulin levels in cattle. J. Anim. Sci. 33:438. 19 McClymont, G. L., and S. Vallance. 1962. Depression of blood glycerides and milk fat synthesis by glucose infusion. Proc. Nutr. Sci. 21 :xli. 20 McCullough, M. E. 1966. Relationships between rumen fluid volatile fatty acids and milk fat percentage and feed intake. J. Dairy Sci. 49:896. 21 Opstvedt, J., R. L. Baldwin, and M. Ronning. 1967. Effect of diet upon activities of several enzymes in abdominal adipose and mammary tissues in the lactating dairy cow. J. Dairy Sci. 50:108.
209
22 Orskov, E. R., R. W. Mayes, and A. Penn. 1971. The capacity for the removal of gtucose from the small intestine by mature sheep. Proc. Nutr. Soc., 30:43a. 23 Palmquist, D. L., C. L. Davis, R. E. Brown, and D. S. Sachan. 1969. Availability and metabolism of various substrates in ruminants. V. Entry rate into the body and incorporation into milk fat of D(-)beta-hydroxybutyrate. J. Dairy Sci. 52:633. 24 Rao, D. R., G. E. Hawkins, and R. C. Smith. 1973. Effect of glucose and insulin on lipoprotein lipase activity in adipose tissue and milk. J. Dairy Sci. 56:1415. 25 Rook, J. A. F., and C. C. Balch. 1961. The effects of intraruminal infusions of acetic, propionic and butyric acids on the yield and composition of the milk of the cows. Br. J. Nutr. 15:361. 26 Rook, J. A. F., C. C. Balch, and V. W. Johnson. 1965. Further observations on the effects of intraruminal infusions of volatile fatty acids and of lactic acid on the yield and composition of the milk of the cow. Br. J. Nutr. 19:93. 27 Satter, L. D., and A. N. Bringe. 1969. Effect of abrupt ration changes on milk and blood components. J. Dairy Sci. 52:1776. 28 Spires, H. R. 1974. Effect of postruminal infusions of glucose and sodium caseinate on milk production, amino acid utilization and glucose synthesis of lactating cows. M. S. Thesis. University of Illinois, Urbana. 29 Storry, J. E., and J. A. F. Rook. 1965. Effects of intravenous infusions of acetate, beta-hydroxybutyrate, triglyceride and other metabolites on the composition of the milk fat and blood in cows. Biochem. J. 97:879. 30 Trenkle, A. 1972. Radioimmunoassay of plasma hormones: Review of plasma insulin in ruminants. J. Dairy Sci. 55:1200. 31 Van Soest, P. J., and N. N. Allen. 1959. Studies on the relationships between rumen acids and fat metabolism of ruminants fed on restricted roughage diets. J. Dairy Sci. 42:1977. 32 Vik-Mo, L., R. S. Emery, and J. T. Huber. 1974. Milk protein production in cows abomasally infused with casein or glucose. J. Dairy Sci. 57:869. 33 Yang, Y. T., and R. L. Baldwin. 1973. Lipolysis in isolated cow adipose cells. J. Dairy Sci. 56: 366.
Journal of Dairy Science Vol. 60, No. 2