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Evaluating Effects of Fish Meal on Milk Fat Yield of Dairy Cows
Evaluating Effects of Fish Meal on Milk Fat Yield of Dairy Cows J. N. SPAIN1 and C. E. POLAN* Department of Dairy Science Virginia Agricultural Experiment Station College of Agrlculture and Life Sciences Virginla Polytechnic Institute and State University Biacksburg 24061-0315 B. A. WATKINS Departments of Food Science and Foods and Nutrition Purdue University West Lafayette, IN 47907-1160 ABSTRACT
Experiment 1 measured the effect of different amounts of dietary fish meal on milk yield and composition. Milk fat percentage and yield were decreased by increased fish meal intake, but this change was not associated with changes in ruminal fermentation patterns. Plasma long-chain n-3 polyunsaturated fatty acids were increased as intake of fish meal increased. Ruminal disappearance of DM, CP, and lipid in fish meal was measured in situ. Seventy percent of lipid disappeared by 8 h. Intraruminal administration of fish oil did not alter ruminal fermentation and only slightly changed fatty acid profiles in duodenal digesta, plasma, or milk. Duodenal infusion increased plasma n-3 fatty acids but did not affect composition of fatty acids in milk. Experiment 2 compared effects of dietary fish meal and fish oil on milk production and composition. Fish meal increased n-3 fatty acids in plasma compared with those of the fish oil treatment. No changes were found in milk yield or composition because of experimental treatments. Cows fed fish meal or fish oil differed significantly in plasma fatty
acid profiles but did not differ in ruminal VFA concentrations or milk fat yield. (Key words: fish meal, milk fat, fatty
acids, yield) Abbreviation key: FM = fish meal, PUFA = polyunsaturated fatty acids. INTRODUCTION
Crude protein recommendations for dairy cows include a requirement for intake of ruminally undegradable protein (17). Several studies (3, 9, 11, 19) have shown improved performance when high quality animal protein products were included in the diets of ruminants. Zerbini et al. (31) found that flow of methionine to the duodenum increased when fish meal (FM) was substituted for soybean meal in diets of lactating dairy cows. Schwab et al. (25) reported that methionine may be a limiting amino acid for milk yield of dairy cows. Therefore, FM has been utilized as an undegradable protein source in diets of lactating dairy cows. Several studies (2, 29) reported increased milk yield from addition of FM to diets of lactating dairy cows. However, in several studies (12, 26, 29), FM supplementation also lowered milk fat content and yields. Currently, several theories have been postulated regarding the influence of FM on milk fat percentage and yields. Given the lipid content of FM, ruminal fiber digestion may be decreased by the presence of polyunsaturated Received January 5, 1994. fatty acids (PUFA) in FM (21). Fish meal Accepted January 6, 1995. contains approximately 10% residual lipid and Uuicnt address: Department of A n i d Science, significant amounts of long-chain n-3 PUFA. University of Missouri, Columbia 6521 1. Varman et al. (28) reported that the inclusion 2To whom reprint request should be made. ~
1995 J L h r y Sci 781142-1153
1142
EFFECTS OF
1143
FISH MEAL ON MILK FAT
of PUFA in diets of lactating cows decreased the molar percentage of ruminal acetate and milk fat percentages, but noted that differences in ruminal VFA patterns were small compared with the large decrease in milk fat, suggesting that ruminal escape of PUFA may have allowed it to alter postruminal lipid metabolism. Infusion of cod liver oil into the rumen or abomasum also decreased milk fat percentage of lactating cows (22). These data support the postabsorptive effect of PUFA on lipid metabolism, as proposed by Vannan et al. (28), which includes decreased uptake of plasma fatty acids by the mammary gland. Effects of marine lipids and, more specifically, long-chain n-3 PUFA on lipid metabolism have been extensively researched. Eicosapentaenoic acid has been shown to inhibit activity of hepatic lipogenic enzymes (6,7). Our laboratory has found that cows fed FM had significantly higher plasma eicosapentaenoic acid and decosahexaenoic acid than These data suggest FM fatty cows not fed M. acids may alter post absorptive lipid metabolism. Therefore, a series of studies was conducted to reevaluate the use of FM as a protein source for lactating dairy cows. In all experi-
ments, a primary goal was to determine site of action by which fish meal has its negative effect on milk fat production. MATERIALS AND METHODS Ewrlmsnt 1
Eight Holstein cows were assigned to two concurrent replicates of a 4 x 4 Latin square, which included four cows per replicate receiving four different diets over four consecutive 20-d periods. Cows averaged 131 f 14 d of lactation and 27 f 2 kg of milk/d at the start of the study. Diets were T M R based on corn silage (Table 1) and were fed twice daily (O600 and 1500 h) in amounts adequate to ensure 10% om. Fish meal was substituted for corn gluten meal on an isonitrogenous basis. Cows were adjusted to diets for a 7-d transition period, followed by a 13-d collection period. Milk yield and DMI were recorded daily. Milk samples were collected at consecutive milkings on d 7 and 13 of the collection period and placed in sealed plastic bags containing potassium dichromate (150 mg per sample) as a preservative. Samples were stored at room
TABLE 1. Ingredient and chemical composition of diets.' Experiment 1 Composition
0% FM
2.6%
FM
5.2% FM
7.8%
FM
Basal diet Experiment 2l In situ Infusion C FM FO ,.- -- ----,
Ingredient Alfalfa silage Barley silage Corn silage thy corn High moisture corn Dried brewers grain soybean meal
(4%) Corn gluten meal FM Mineral mix Limestone chemical
DM, % ADF CP
... ...
... ...
... ...
...
62.3
62.9
62.0
61.9
...
...
35.8
...
...
36.1
77.9
37.5
...
37.6
8.2
51.9 16.6
...
...
...
16.4
...
35.8
... 37.3
...
...
...
20.3
20.5
20.2
20.2
...
...
14.9
15.0
14.9
...
...
...
...
...
3.6
...
...
...
10.1 7.3
10.3 3.7 2.6
10.1 2.5
...
12.2
7.8 2.9
7.5 3.7
...
7.5
7.5 3.9
...
... ...
42.0 25.2 17.2
5.2
...
10.1 7.8
...
...
...
1.7
.8
...
... ...
...
...
...
...
44.0 25.1 17.5
44.0 24.7 17.3
42.0 24.4 17.2
48.9 24.2 17.4
...
...
19.4 17.5
... ...
3.8
...
.6
...
44.3 24.5 15.7
44.0 24.2 16.3
...
. . . .6 44.3 24.2 15.5
IC = Control, FM = fish meal, and FO = fish oil, Journal of Dairy Science Vol. 78. No. 5, 1995
1144
SPAIN ET AL.
temperature for 24 h before being analyzed with a four-channel spectrophotometer (Multispec Mark I; Foss Food Technology, Eden Prairie, MN) for milk fat, protein, and lactose by Virginia DHIA. Approximately 150 ml of ruminal fluid were sampled by aspiration via esophogeal tube 2 to 3 h after the p.m. feeding on d 12 of the collection period. A 5-ml aliquot was placed in a plastic tube containing four drops of concentrated sulfuric acid for ammonia analysis. Another 10-ml aliquot was stored frozen (-2O'C) in a separate plastic tube for VFA analysis. Also, on d 12 of the collection period, 30 ml of blood were sampled by jugular puncture. Ten-milliliter subsamples were placed in plastic tubes containing 100 units of heparin each. All samples were stored on ice for transport to the laboratory. Blood samples were centrifuged at 3220 x g for 20 min at 5'C. Plasma was decanted and stored at -2O'C until analyzed. Plasma used for fatty acid analysis was stored at -8o'C. Ruminal fluid was stored at -2O'C until analyzed. Plasma urea was determined colorimetrically (4). Samples of diet components and TMR were collected weekly, and DM was determined by drying samples at 1WC in a forced-air oven for 24 h. Dried samples were ground to pass through a 2-mm screen (Wiley mill; Arthur H. Thomas, Philadelphia, PA). Kjeldahl N (1) and ADF (5) were measured on ground subsamples. Plasma lipids were extracted with ch1oroform:methanol (2: 1; volhol). Methyl esters of fatty acids from plasma, menhaden oil, duodenal ingesta, and milk fat were prepared by using 14% boron trifluoride and analyzed by capillary GLC (23). The methyl esters were extracted in hexane for chromatographic analysis by an HP 5890A gas chromatograph equipped with a flame ionization detector and integrator (Hewlett-Packard Co., Avondale, PA). A DM 225 (25% cyanopropylphenyl) fused silica capillary column, 30 m x .25 mm i.d. (J & W Scientific Co., Rancho Cordova, CA), was used with helium as the carrier gas. The initial oven temperature of 196'C was held for 12 min and increased at a rate of .9"U min until the final temperature of 214°C was reached. The total GLC run time was 40 min. An external standard mixture prepared from known amounts of triacylglycerols and methylated fatty acids (Nu Check-Prep, ElyJournal of Dairy Science Vol. 78, No. 5, 1995
sian, MN) was used to obtain retention times and to develop the calibration table. Heptadecaenoic acid (17: 1, voVvol) was added to the external standard mixture and to all samples, except the duodenal digesta, as the internal standard. Fatty acid values are presented as micrograms per milligram in the menhaden oil and milk fat and as micrograms per milliliter of plasma. Fatty acid values in duodenal digesta are reported as area percentages. Acidified ruminal fluid samples were thawed at room temperature, centrifuged at 3220 x g for 20 min, and analyzed for ammonia (13). Ruminal VFA concentrations were determined by the method previously used by Spain et al. (26). Data were analyzed statistically using the general linear models procedure of SAS (16). The statistical model used was Y = p + Ci + Trt, + Pdk + (TrtPd),k, where p is the population mean, C, is the cow i., Trt, is treatment j, Pdk is period k, and (TrtPd)jk. is the interaction between treatment j and penod k (10). Treatment means were tested by error means squares. Differences among means were evaluated by Tukey's procedure (10). Linear and quadratic response relationships were measured by orthogonal contrasts. In Situ Characterization of FM
Three ruminally fistulated cows were housed in a tie-stall barn and were individually fed a TMR (Table 1) twice daily. Approximately 10 g of FM were placed in dacron polyester bags (143 cm2) with a pore size of 57 pm. Bags were fastened to metal chains attached to nylon cords with three bags per incubation period per cow. Ruminal resident times for the bags were 2, 4, 8, 12, 24, 48, 72, and 120 h. At the beginning of incubation, bags were placed on the ventral floor of the rumen, and they were allowed to mix and be moved within the rumen. Bags were placed in the rumen in reverse order over time to allow the simultaneous removal of all bags. Upon removal, bags were individually rinsed to remove particles from the bag exterior. Bags were then placed in a continuous tap water rinse system for 24 h. During the continuous rinse, the system was emptied twice and refilled with fresh water. After rinsing, bags were suspended in a forced-air oven
1145
EFFECTS OF FISH MEAL ON MILK FAT
and dried for 48 h at 65°C. Dried residues were composited for each cow by residence time period. Composites were ground through a .5mm screen. Each composite was analyzed for DM and CP (1) as previously described. Lipid was analyzed by refluxing residue with petroleum ether for 1 h, and solvent was then removed (Soxtec System HT; Tecator, Herndon, VA). The remaining lipid was then weighed. Marlne Lipid infusion
Six lactating Holsteins (130 f 9 d of lactation) were assigned to two concurrent replicates of a 3 x 3 Latin square. The design included three 10-d periods during which cows received one of three treatments. Cows had been surgically fitted with ruminal fistulas and duodenal T-type cannulas. At the initiation of experiment, mean milk yield was 34.0 f 3.9 kg/d. Cows were housed in a tie-stall barn. Treatments were corn gluten meal administered into the rumen, corn gluten meal plus fish oil administered into the rumen, and corn gluten meal administered into the rumen plus fish oil infused into the duodenum. All cows were fed the same basal diet twice daily for 7 d prior to initiation of and throughout the 30-d experiment (Table 1). Cows were fed 30% of their daily ration at O600 h and the remaining 70% at 1500 h. Orts were removed daily and sampled for individual cows every 3 d of each period. The corn gluten meal was a mixture (86:14, d w t ) of corn gluten meal and limestone. The 341-g dose was placed into the rumen twice daily. The amount given was equivalent to the protein, calcium, and, when appropriate, fish oil in an equal amount of FM. Fish oil was a crude menhaden oil, and cows were dosed twice daily (48 d d ) 2 h after each feeding. Cows receiving the duodenal infusion of fish oil received one-half of the dose 2 h after feeding with the remainder given 1 h later. The fatty acid composition of the crude fish oil is shown in Table 2. Cows were milked twice daily (1200 and 2400 h), and yield was recorded. Milk samples were collected at consecutive a.m. and p.m. milkings on d 4.7, and 10 of each experimental period. Samples were handled and analyzed as described for Experiment 1. On d 10, 1 L of milk was collected at the p.m. milking and
TABLE 2. Fatty acid composition of crude Menhaden fish Oil'.
Fatty acid
Wmg)
c12:o c14:O
Trace 85 5 175 100 12 7 30
C18:l C18:l c18:2 c18:2 C18:3n6 C18:3n6 C18:3n3 C18:3n3 C Cm m 00
Cm1 Cm1
c20:2 c20:2
c20:3n6 c20:3n6 CD4n6 CD4n6 C20:3n3 C20:3n3 C205n3 C205n3 c22: c22: 1 1n9 n9 c22:6n3 C22.hn3
64 48 2 7 3 8 Trace 10 Trace Trace 112 Trace
59
IMenhaden fish oil was an unrefined, crude oil that was used to compare with residual crude oil associated with fish meal.
transported to the laboratory at ambient temperature. A 225-m1 aliquot was centrifuged at 9500 x g for 1 h at 10°C. The lower aqueous layer was aspirated by suction to isolate the fat layer. The milk fat was placed in individual plastic bags, which were sealed in larger plastic bags and stored at -7O'C until fatty acid analysis. Samples of ruminal fluid and blood were collected 2 h postdosing on d 10. Ruminal fluid was collected from various regions of the rumen and filtered through four layers of cheesecloth. Ruminal pH was measured immediately upon collection of the fluid. Ruminal fluid was mixed and subsampled for ammonia and VFA analysis, as outlined for Experiment 1. Blood samples were collected by puncture of the jugular vein, transferred to plastic tubes containing 200U of heparin, and processed as described for Experiment 1. Plasma was decanted and stored in plastic tubes at -20°C until analyzed. Plasma samples used for fatty acid analysis were stored at -70°C until analysis, as previously described. One liter of continuous duodenal digesta samples was collected at 0100, 0500, 0900, Journal of Dairy Science Vol. 78, No. 5, 1995
1146
SPAIN ET AL.
1300, 1700, and 2100 h over a 3-d period, representing 4-h intervals in a 24-h day. Duodenal digesta pH was measured electronically using a pH meter. Duodenal samples were stored in sealed plastic cups at -2O'C. For analysis, duodenal samples were thawed to room temperature and composited by cow for each period. Dry matter content was determined as the loss of weight following freezedrying for 120 h. Dried samples were ground to pass through a .5-mm screen. Acid detergent fiber was measured by the method of Goering and Van Soest (5). Acid-insoluble ash was determined by ashing the ADF residue at 500'C for 4 h. Acid-insoluble ash was a reference marker used to calculate postruminal flow. Fatty acids of duodenal DM were extracted by the method of Outen et al. (20) and analyzed by GLC. Duodenal cytosine was determined as described by Zerbini et al. (31). Diet component samples were collected weekly. The ADF and acid detergent-insoluble ash were measured as described for duodenal samples. Kjeldahl N was measured and reported as CP. The experimental model was Y = p + replicate + cow(replicate) + period + treatment, where p = treatments were tested by the error means square, and means were compared by Tukey's procedure (15). Statistical analysis was by the general linear models procedure of SAS
twice daily, and yield was recorded at each milking. Milk samples were collected at four consecutive milkings on d 16, 17, and 18 and analyzed for fat, protein, and lactose as described previously. Blood and ruminal samples were taken 2 h following the morning feeding and analyzed for plasma urea N, plasma long-chain fatty acids, ruminal ammonia, and ruminal VFA using procedures reported earlier. Dietary treatments were arranged in a 3 x 3 Latin square design with four replicates conducted concurrently. The experimental model was defined as Y = p + replicate + cow(rep1icate.) + period + treatment + residual with treatment means squares tested by residual means squares (10). Means were separated by Tukey's procedure (15). Statistical analysis was by the general linear models procedure of SAS
(W. RESULTS AND DISCUSSION Experiment 1
In Experiment 1, DMI was greatest when diets contained 2.6% FM and was lowest when 7.8% FM was included in the diet, resulting in a quadratic response (Table 3). Although inclusion of FM significantly affected DMI, the differences in intake were not of sufficient magnitude to alter milk production. Milk fat (W. percentage decreased linearly as intake of FM increased from 3.5% for controls to 3.0% for Experiment 2 cows fed the 7.8% FM diet. Milk protein and Twelve lactating Holstein cows (mean lacta- lactose percentages were not affected by the tion, 3.1) were assigned to one of four repli- diet. cates of a 3 x 3 Latin square design. Cows Ruminal N H 3 N appeared to respond in a were housed in a tie-stall barn and fed one of quadratic manner (P < .15); the value was three diets over three 18-d periods. Diets were highest (6.6 mgldl) when 2.6% FM was fed, T M R based on corn silage and alfalfa haylage and lowest (4.8 mg/dl) when 0 or 7.8% FM (Table 1). Fish oil was mixed with a corn was fed. Treatment means paralleled the patgluten meal and mineral premix, placed in tern for DMI (P < .15). Concentrations of plastic bags, and stored at 6'C. This premix plasma glucose (70 mgldl) and urea (21 mddl) contained corn gluten meal (80.0%), fish oil were also similar across the diets. (7.2%), and limestone (12.8%), making the Differences in milk fat cannot be attributed content of fish oil and calcium in the mix to altered ruminal fermentation. Total concensimilar to that in FM. Ethoxyquin (500 ppm) trations of VFA in ruminal fluid ranged from was added as an antioxidant during premixing 145.3 to 165.6 mM but did not differ with diet. of the fish oil with the corn gluten meal plus Individual VFA concentrations also were unlimestone. Cows were fed twice daily at O600 affected by diets. The molar percentage of and 1500 h in amounts to ensure 10% orts. butyrate was higher (P < .05; 13.5 vs. 12.5 Orts were recorded 4 dwk. Cows were milked moVl00 mol) for cows fed 5.2% FM than for Journal of Dairy Science Vol. 78, No. 5, 1995
1147
EFFECTS OF FISH MEAL ON MILK FAT TABLE 3. Effect of fish meal (FM) on DMI. milk yield, and milk composition in Experiment 1. Dietary treatments Item
0% FM
2.6% FM
5.2% FM
7.8% FM
SEM
DM1.l kg/d Milk, kg/d Fat,z % Protein, 96 Lactose. %
20.w 25.3 3.4
21.21 26.2 3.2ab 3.4
20.1b 25.5 3.1b 3.4
19Sb 25.6 3.P 3.3
.1 .4 .04 .01
4.1
4.1
47
47
3.53
apbMeans in same row followed by different superscripts differ (P < .05). lsignificant quadratic response to dietary treatment. ZSignificant linear response to dietary treatment.
cows fed 7.8% FM. The acetate to propionate ratio ranged from 2.6 to 2.8 but was not different among treatments. Jenny et al. (8) reported that decreased milk fat was associated with shifts in ruminal VFA patterns and concurrent increase in concentrations of plasma glucose. Plasma glucose was highest for cows fed high FM diet (73.6 mg/dl) but was not different from means of other treatments. Cows fed linear increments of FM exhibited a concurrent linear decrease in milk fat content in the absence of significant changes in milk volume, ruminal VFA patterns, or plasma glucose. Inclusion of FM in increasing concentrations did alter plasma PUFA profiles (Table 4).
TABLE 4. Effect of different amounts of fish meal increasing amounts of FM in Experiment 1.
Unsaturated fatty acids C16:ln7, C18:3n3, and C22:6n3 increased linearly (Pc .05) as FM intake increased. In contrast, C18:3n6 and C20:3n6 decreased linearly ( p < .05) as FM intake increased. Saturated fatty acids and the fatty acids C18:ln9 and C18:2n6 were not altered by the diet. It is important to note that C20:5n3 and C22:6n3 survived microbial biohydrogenation, thus allowing for the absorption of these long-chain PUFA. With regard to metabolic impact of these n-3 PUFA, reports (6, 7, 30)indicate that inclusion of n-3 PUFA in experimental diets of nonruminants resulted in a decreased activity of several key lipogenic enzymes. Activities of acetyl coenzyme A car-
C204n6, C20:5n3,
on plasma concentrationsof long-chain fatty acid of cows fed Diet
Fatty acid
0% FM
2.6% FM
5.2% FM
7.8% Fhl
SEM
237 19b 19 359 167 622
38 1 2 I89 18 21 1 1 1 3 2
~~
c16:O C16:1A c17:O C18 C18:ln9 C18:2n6 c18:3n6A C18:3n3A C203116~ CU):4n6A C205dA c224n6 C226n3A
239 12' 18 384 126 708 16 22a 436 38 91 5 9
otg/ml) 187 14l 15 292 117 713 11 27.b 35.b 47 30h 5 17
198
123 16 318 121 690 14 233 4o.b
41 19*b 5 13 ~~
10
3Ib 31b 51
42C 4 22
1
2
__
l.b~cMeansin same row followed by different letters differ (P < .05). ASignificant linear effects to diet (P < .Os).
Journal of Dairy Science Vol. 78, No. 5, 1995
1148
SPAIN ET AL.
TABLE 5. Disappearance1 of DM, CP. and ether extract of fish meal during in situ incubation in the rumen.
0 2 4 8 12 24 48 72 120
(%)
28.9 29.8 31.4 32.5 37.1 50.2 59.8 71.8
17.6 17.9 22.5 22.0 30.6 49.7 64.4 82.8
75.2 84.2 83.9 84.4 92.1 92.1 88.9 92.9
Percentage disappearance = 100 minus the percentage remaining in in situ bag at a given time.
ble 5). These results were similar to those reported by Zerbini et al. (31). In contrast to DM and CP, ether extract was removed very rapidly; 75% was removed after 2 h, and 92% of original lipid was removed after 24 h. Obviously, fat in FM is readily a part of the ruminal environment and may affect microbial metabolism in the rumen (17). These data do not address the availability of specific fatty acids, which may differ in their release and uptake. Individual fatty acid residues may not alter total flow to postruminal absorptive sites. The potential of the n-3 PUFA to alter lipid metabolism makes this lipid material especially important to evaluate. Firh 01 inturion Trial
The DMI averaged 19.7 kg/d and was not different among treatments (Table 6). Daily milk yields also were similar among treatments and averaged 31.6 kg/d. No differences occurred in milk composition and component yields among treatments. Ruminal VFA patterns were also similar across diets (Table 7). 2). Although total VFA concentration tended to be lower for ruminal infusion of fish oil, the in Situ Trial differences were not significant. Pennington Data from Experiment 1 suggested that marine PUFA passed from the rumen; therefore, and Davis (22) decreased milk fat content by the in situ experiment was conducted to evalu- infusing 225 g of destearinated cod liver oil ate ruminal degradation characteristics of men- into the rumen or abomasum. The change in haden FM. Preincubated Fh4 contained 94.3% milk composition occurred without changes in DM, 61.8% CP, and 9.4% fat, which were ruminal VFA patterns. Varman et al. (28) similar to previously reported values (9). By 24 reported similar results from dietary fish oil h, 37 and 30.6% of the original DM and CP, supplement fed to lactating cows. Ruminal respectively, were removed from the bag (Ta- acetate decreased slightly by inclusion of fish boxylase and fatty acid synthetase were greatly reduced for mice fed diets high in eicosapentaenoic acid (C205n3). This fatty acid is also found in plasma of mice fed marine oil containing high amounts of C205n3 (6). Up to 25% of the fatty acids in FM are n-3 PUFA (Table
TABLE 6. Effect of ruminal or duodenal infusion of fish oil on intake, milk yield, composition, and component yields. Infusion method Item
None
Ruminal
DMI Milk Fat Protein Lactose
19.7 31.3
20.2 31.5 1 .o 1 .o 1.5
Duodenal
SEM
19.2 32.0 1 .o 1 .o 1.5
.2 .2
3.2 3.1
3.1 3.1
.04 .01
4.8
4.1
.01
Wd)
I .o
.o
1 1.5
.o 1 .01 .o1
(W Fat Protein Lactose
3.2 3.1 4.7
Journal of Dairy Science Vol. 78, No. 5, 1995
1149
EFFECTS OF FISH MEAL ON MILK FAT
TABLE 7. Effect of site of infusion of fish oil in the rumen or duodenum on ruminal VFA concentrations, ratios, and pH. lnfusion method Item
None
Ruminal
Total Acetate (A) Propionate (P) Isobutyrate Butyrate Isovalerate Valerate A:P Ruminal pH
150.9 82.0 40.2 1.6 20.9 3.1 3.2 2.2 5.9
140.1 80.0 33.5 1.6 18.7 3.1 3.4 2.5 5.8
Duodenal
SEM
150.7 86.8 34.7 1.7 20.5 3.5 3.6 2.7 6.0
2.1 1.6 2.0 .03 .1 .1 .1 .2 .02
(mM)
oil but not of the magnitude associated with a large decrease in milk fat content that they observed. V m a n et al. (28) reported that arteriovenous differences in mammary triglyceride uptake decreased, suggesting a possible postabsorptive effect of fish oil fatty acids. In contrast, Storry et al. (27) reported decreased ruminal acetate in cows fed 300 g of unprotected cod liver oil, which resulted in decreased milk fat production. In all of these reports, fish oil amounts were greater than those fed in this experiment (-50 gld) and in Experiment 1 (-150 g/d). In Experiment 1, corn silage was the only forage, but alfalfa provided 16 and 35% of the DM in the infusion experiment and Experiment 2, which may have resulted in less milk fat response, as observed by Broderick (3). Duodenal digesta flows (14.5 kgld) were not different among treatments. Duodenal digesta pH and cytosine concentration were also unaffected by treatment. These results, in addition to the lack of differences in DMI and milk yield, further support the conclusion that fish oil did not alter ruminal fermentation in the present study. Although small differences were measured in fatty acid contents of duodenal digesta, these differences would be expected to have little metabolic consequence for the animal (Table 8); however, infusion of fish oil altered plasma fatty acid concentrations, and the response differed among treatments (Table 9). Saturated fatty acids and the monoenoic Cl8:ln9 were not different because of treatment. Palmitoleic acid (C16:ln7) was increased
(P-c .09) by duodenal infusion of fish oil. This small difference was probably of little consequence to the cow; however, PUFA were markedly changed by infusion of fish oil. yLinolenic (cl8:3n6) was decreased by infusion of fish oil compared with that of control cows. The COnCen@i3tiOnSOf C204n6, C20:5n3. and c 2 2 6 f l were all increased by duodenal infusion of fish oil. Apparently, because of microbial metabolism in the rumen, administration of fish oil into the rumen failed to cause changes in plasma fatty acid profiles. These differences associated with the source of oil between Experiment 1 and the infusion study suggest differences in ruminal stability or microbial metabolism of fat in fish oil versus FM. Although plasma fatty acids were altered, milk fatty acid composition did not differ from C 4 to c l 8 including C18:Ov C 1 8 : l ~ c 1 8 : 2 ~ C18:3n6, and C18:3n3. Only trace amounts above c 1 8 were found. Studies in which whole oil seeds or protected lipids have been fed have shown significantly altered fatty acid composition of milk (13, 16). Pennington and Davis (22) increased unsaturated fatty acid composition of milk with infusion of fish oil. Nicholson and Sutton (18) reported a linear increase of c16:1 in milk fat as fish oil increased. Pennington and Davis (22) also reported increases in the presence of unidentified 20-carbon fatty acids after cows were infused with fish oil. Dosage in the present study was lower than those previously reported but was calculated to reflect normal lipid intake from a diet supplemented with FM. This lack of reJournal of Dairy Science Vol. 78. No. 5, 1995
1150
SPAIN ET AL.
TABLE 8. Effect of site of infusion of tish oil on fatty acid composition of duodenal digesta. Infusion method Fatty acid c4:0 c6:0 C8:O
c1o:o c120 c13:0 c140 c141
c15:o C1S:l c16:0
c16:I c17:0
Cl8:O Cl8:l Cl8:2 I8:3n6 C18:3n3
c20:o
None .8* .2 .4
.2 .4 .1 1.5 .2 1.o .5 18.3 .3 1.1 51.8 10.8 7.3
Trace .7 .8a
Ruminal
Duodenal
.4b .2 .4 .2 .4 .2 I .7 .2 1.o .4
18.9 .3 1.2 48.7 14.4 7.7 TraCe .E 1.Ob
SEM
.5* .2
.05 .01
.4
.02
.2 .4 .2 1.4 .2 1.o .5 17.8 .2 1.1 45.5 11.3 10.4 TraCe
.02 .01 .02 .04
.8 .Ea
.02
.01 .05 .2 .03 .02 1.o 1.o .5
.02 .02
*.bMeans in same row with different superscripts differ (P < .05).
sponse, compared with previously reported studies (14), may also result from the preferential use of saturated and monoenoic fatty acids for milk triglyceride synthesis; furthermore, length of experimental periods might have been inadequate to allow equilibration of fatty acids and maximum metabolic responses to changes in plasma fatty acid concentrations.
Neither ruminal total VFA concentrations (96.4 mM) nor individual VFA differed with diet. Acetate:propionate ratios averaged 3.4.
This lack of response from the inclusion of FM or fish oil agrees with published results (22). Nicholson and Sutton (18) have reported that the addition of cod liver oil to diets of dauy cows reduced ruminal acetate and butyrate concentrations; however, fiber influenced VFA concentrations, and the change was more proExperiment 2 nounced for the low roughage diets than for With the apparent differences in digestion medium or high roughage diets. Fiber source between FM and fish oil, this experiment was may also influence response to M or fish oil. conducted to compare feeding free oil or oil Broderick (3) reported FM addition to diets indigenous in FM. Diet did not affect DMI or based on alfalfa silage did not change milk milk yield (Table lo), which agrees with previ- composition. Alfalfa supplied 36% of the DM ous experiments showing no differences on in Experiment 2. Differences because of yield when FM was substituted for corn gluten forages may reflect differences in ruminal fermeal. Milk fat decreased .1 percentage units by mentation. Differences in microbial fermentaFM and fish oil, but total fat yield was equal. tion in the rumen, caused by soluble and strucNo other differences in component concentra- tural carbohydrates, may cause hormonal tions or yields were measured. When Oldham changes in the animal and alter fatty acid et al. (19) substituted FM for urea in diets of metabolism (8), which may change the effect lactating dairy cows, milk protein yields were of n-3 fatty acids on lipid metabolism. significantly increased. Substitution of FM for Shifts in plasma fatty acid concentrations soybean oil meal did not alter milk composi- were similar to those in the previous two tion. studies (Table 11). The FM tended to lower Journal of Dairy Science Vol. 78, No. 5, 1995
1151
EFFECTS OF FISH MEAL ON MILK FAT TABLE 9. Effect of site of fish oil infusion on long-chain fatty acid composition in plasma. Infusion Fatty acid
None
C160
28 1 14 24 443 181 1loo 301 66 69a 17a 8 20 Trace.
Ruminal
Duodenal
SEM
OCW) C16I c17:0 c18:0 c18:1 c18:2 C18:3n6 C20:3n6 C204n6 C205n3 C22:4n6 C22:5n3 C22:6n3
258 15 24
294 24 25
409
440
193 1163 25b 69 701 2ga 9 19 2'
174 1126 22b 65 82b 64b
a*bMeans in same row with different superscripts differ
C18:2n6$ C18:3n6. and C20:3n6. In contrast, C16:1, Cl8:3n3, C20:4n6*.C20:5n39 and C22:6n3 were in-
creased by the inclusion of FM in the diet. Fish oil increased only C20:5n3 when compared with fatty acids of the control diet. These responses agree with the previous work done at our lab with a similar percentage of FM. These results indicate a difference exists in the ruminal characteristic between the FM and fish oil as sources of supplemental marine lipids; however, in previous experiments, FM addition to diets based on corn silage was accompanied by decreased milk fat content (26). In the present study, milk fat decreased only slightly with a similar increase in plasma n-3
20 2 2 30 11 18 1 2 2 1
10
1
28 12b
2 1
(P c .05).
fatty acids. Potential for forage source to influence yield responses should be considered (3). CONCLUSIONS
Fish meal altered milk fat percentage when fed in diets based on corn silage. This change in milk composition was not associated with changes in patterns of ruminal VFA. Increased concentrations of n-3 fatty acids were measured with the inclusion of FM, suggesting a protection of these PUFA from microbial metabolism in the rumen. Use of fish oil as a comparison for FM seemed to produce a
TABLE 10. Effect of dietary treatment on milk yield, DMI, milk composition, and component yields of cows on Experiment 2. Diet
Ikm
Control
Fish meal
Fish oil
SEM
Milk, kg/d
30.1 21.6
30.3 21.2
30.1 21.4
.2 .3
3.8 1.1
3.7 1.1
3.7 1.1
.03 .01
3.1 .9
3. I .9
3.1 .9
.01
4.8 1.4
4.8 1.4
4.8 1.4
.01
DM, kg/d Milk fat 5%
kdd Milk protein 5%
Wd Milk lactose 5%
kg/d
.01
.01
Journal of Dairy Science Vol. 78, No. 5, 1995
1152
SPAIN ET AL
TABLE 11. Effect of dietary fish meal or fish oil fed to lactating cows on fatty acid concentrations in plasma of cows in Experiment 2. Diet
Item
Control
c16:0 c16:1 c17:0
268 19. 23 397 172 1331 31 142 74 7 9 26' 9 22 4'
Fish meal
Fish oil
SEM
286 26b 24 391 182 1195 23 167 65 87b 60b 12 27 18b
8 .7 .7 12
Wm)
C18:O
C18:l C18:2 C18:3n6 18:3n3 C20:3n6 C20:4n6 C205n3 C22:4n6 C225n3
C226n3
288 2or 24 419 175 1334 29 146 74 723 32b 11 22 4'
5
42 1 4 2 2 2 .7 1 1
aJ'Means in same row followed by different a superscript differ (P < .05).
different response based on changes in plasma fatty acid profiles. The differences in the plasma fatty acid response between ruminal infusion or dietary fish oil, compared with duodenal infusion of fish and dietary FM,support this observation. The exact role of n-3 PUFA on milk lipid yield should be investigated further. At sufficiently high amounts of dietary fatty acids from fish, ruminal activity would probably be altered, as has been observed with other unsaturated fatty acids. However, VFA patterns were not altered in this study, even though milk fat declined. Fish oil is released readily from FM.Perhaps amounts infused or fed as FM were too small to elicit a response, even though the amounts fed coincided with practical feeding amounts. The inclusion of alfalfa in the diet may also have influenced the response. The ability to manipulate milk fat yield without excessive grain feeding would be attractive, given current changes in milk fat demand. The interaction of forage base and response to n-3 PUFA still needs investigation. REFERENCES 1 Association of Official Analytical Chemists. 1980. Official Methods of Analysis. 13th ed. AOAC, Washington, DC. 2Atwal. A. S., and J. D. M e . 1992. Effects of fish mal feeding to cows on digestibility, milk production, and milk composition. J. Dairy Sci. 75502.
Journal of Dairy Science Vol. 78, No. 5 , 1995
3Broderick. G. A. 1992. Relative value of fish meal versus solvent soybean meal for lactating dairy cows fed alfalfa silage as sole forage. J. Dairy Sci. 75:174. 4 Coulombe, J. J., and L. Favreau. 1963. A new simple semimicro method for calorimetric determination of urea. clin. Chcm. 9:102. SGoeMg, H. K., and P. J. Van Soest. 1970. Forage Fiber Analyses (Apparatus, Reagents, Procedures, and some Applications). Agric. Handbook No. 379. ARSUSDA, Washington, DC. 6Huzberg. G. R.. and M. Rogerson. 1988. Hepatic fatty acid synthesis and triglyceride secretion in rats fed fructose or glucose-based diets containing corn oil, tallow or marine oil. J. Nutr. 118:1061. 7 Ifitani, N., K.Inoguchi, M. Endo, E. Fudude, and M. Morita. 1980. Identification of shellfish fatty acids and their effects on lipogenic enzymes. Biochem. Biophys. Acta 68:378. 8 J e ~ y B. . F., C. E. Polan, and F. W. Thye. 1975. Effects of high grain feeding and stage of lactation on Serum insulin, glucose, and milk fat percentage in lactating cows. J. Nutr. 104:379. 9 Klopfenstein, T. 1991. W i n g animal protein products. Page 60 in Pmc. Alternative Feeds for Dairy and Beef Cattle Symp., St. Louis, MO. E. R. Jordon. ed. Sponsored by USDA-Extension Service, industry support, and Univ. Missouri Ext., Columbia. IOLcntner, M., and T. Bishop. 1986. Experimental Design and Analysis. Valley Book Co., Blacksburg, VA. 11 Mbtysaari, P. E.,C. J. Sniffen, T. V. Muscato. J. M. Lynch, and D. M. Barbano. 1989. Performance of cows in early lactation fed isonitrogenous diets containing soybean meal or animal by-product meals. J. Dairy Sci. 72:2958. 12 Mattos, W., and D. L. Palmquist. 1974. Increased polyunsaturated fatty acid yields in milk of cows fed protected fats. J. Dairy Sci. 57:1050.
EFFECTS OF FISH MEAL ON MILK FAT 13 McCullough, H. 1967. The determination of ammonia in whole blood by direct colorimetric method. Clin. Chem. Acta 17:297. 14 Moore, J. H., and W.W.Christie. 1980. Lipid metabolism in the mammary gland of ruminants. Rog. Lipids Res. 17:347. 15 Mosteller, F.. and J. W. Tukey. 1977. Data Analysis and Regression. Addison-Wiley, New York, NY. 16 Murphy, J. J., and D. J. Morgan. 1978. Effect of inclusion of protected and unprotected tallow in dairy rations on cow performance. A n i . Prod. 32:368. 17National Research Council. 1989. Nutrient Requirements of Dairy Cattle. 6th rev. d.Natl. Acad. Sci., Washington, DC. 18Nicholson. J.W.G..and J. D. Sutton. 1971. Some effects of unsaturated oils given to dairy cows with rations of different roughage to concentrate. J. Dairy Res. 38:363. 19Oldham. J. D., D. J. Napper, T. Smith, and R. J. Fulford. 1985. Performance of dairy cows offered isonitrogenous diets containing urea or fish meal in early and mid-lactation. Br. J. Nutr. 53:337. 20Outen. G. E.,D. E. Beever. and 1. S. Fenlon. 1976. Direct methylation of long-chain fatty acids in feeds, digesta, and faeces without prior extraction. J. Sci. Food Agric. 27:419. 21 Palmquist. D. L., and T. C. Jenkii. 1980. Fats in lactation rations: review. J. Dairy Sci. 63:l. 22 Pennington, J. A., and C. L. Davis. 1975. Effects of intraruminal and intra-abomasaladditions of cod-liver on milk fat production in the cow. J. Dairy Sci. 58:49. 23 Phetteplace, H. W., and B. A. Watkins. 1990. Lipid measurements in chickens fed different combinations
1153
of chick fat and menhaden oil. J. Agric. Food Chem. 38:1848. 24 SAS@ User’s Guide: Statistics, Version 5 Edition. 1985. SAS Inst., Inc., C q , NC. 25Schwab. C. G.. C. K. Bo&. N. L. Whitehouse. and M.M.A. Mesbah. 1993. Amino acid limitation and flow to the duodenum at four stages of lactation. 1. Sequence of lysine and methionine limitation. J. Drury Sci. 75:3486. 26Spain, J. N., M. D. Alvarado, C. E. Polan. C. N. Miller, and M. L. McGilliard. 1990. Effect of protein source and energy on milk composition in midlactation dairy cows. J. Dairy Sci. 73:445. 27 Stony, J. E., P. E. Brumby. A. J. Hall, and B. Tuckley. 1974. Effects of free and protected forms of codliver oil on milk fat secretion in the dairy cow. J. Dairy Sci. 57:1046. 28 Varman, P. N., L. H. Schultz, and R. E. Nichols. 1968. Effect of unsaturated oils on rumen fermentation, blood components, and milk composition. J. Dairy Sci. 51:1956. 29 Wohlt, J. E., S. L. Chmiel. P. K.Zajac, L. Backer. D. B. Blethen, and J. L. Evans. 1991. Dry matter intake, milk yield and composition, and nitrogen use in Holstein cows fed soybean, fish. or corn gluten meals. J. Dauy Sci. 74:1609. 30Yang. Y.T., and M. A. Williams, 1978. Comparison of CIS-. Cm-, C 2 + ~ t u m t e dfatty acids in reducing fatty acid synthesis in isolated rat hepatocytes. Biochim. Biophys. Acta 531:133. 31 Zerbini, E., C. E. Polan, and J. H. Herbein. 1988. Effect of dietary soybean meal and fish meal on protein digesta flow in Holstein cows during early and midlactation. J. Dairy Sci. 71:1248.
Journal of Dairy Science Vol. 78, No. 5, 1995
Experiment 1 measured the effect of different amounts of dietary fish meal on milk yield and composition. Milk fat percentage and yield were decreased by increased fish meal intake, but this change was not associated with changes in ruminal fermentation patterns. Plasma long-chain n-3 polyunsaturated fatty acids were increased as intake of fish meal increased. Ruminal disappearance of DM, CP, and lipid in fish meal was measured in situ. Seventy percent of lipid disappeared by 8 h. Intraruminal administration of fish oil did not alter ruminal fermentation and only slightly changed fatty acid profiles in duodenal digesta, plasma, or milk. Duodenal infusion increased plasma n-3 fatty acids but did not affect composition of fatty acids in milk. Experiment 2 compared effects of dietary fish meal and fish oil on milk production and composition. Fish meal increased n-3 fatty acids in plasma compared with those of the fish oil treatment. No changes were found in milk yield or composition because of experimental treatments. Cows fed fish meal or fish oil differed significantly in plasma fatty
acid profiles but did not differ in ruminal VFA concentrations or milk fat yield. (Key words: fish meal, milk fat, fatty
acids, yield) Abbreviation key: FM = fish meal, PUFA = polyunsaturated fatty acids. INTRODUCTION
Crude protein recommendations for dairy cows include a requirement for intake of ruminally undegradable protein (17). Several studies (3, 9, 11, 19) have shown improved performance when high quality animal protein products were included in the diets of ruminants. Zerbini et al. (31) found that flow of methionine to the duodenum increased when fish meal (FM) was substituted for soybean meal in diets of lactating dairy cows. Schwab et al. (25) reported that methionine may be a limiting amino acid for milk yield of dairy cows. Therefore, FM has been utilized as an undegradable protein source in diets of lactating dairy cows. Several studies (2, 29) reported increased milk yield from addition of FM to diets of lactating dairy cows. However, in several studies (12, 26, 29), FM supplementation also lowered milk fat content and yields. Currently, several theories have been postulated regarding the influence of FM on milk fat percentage and yields. Given the lipid content of FM, ruminal fiber digestion may be decreased by the presence of polyunsaturated Received January 5, 1994. fatty acids (PUFA) in FM (21). Fish meal Accepted January 6, 1995. contains approximately 10% residual lipid and Uuicnt address: Department of A n i d Science, significant amounts of long-chain n-3 PUFA. University of Missouri, Columbia 6521 1. Varman et al. (28) reported that the inclusion 2To whom reprint request should be made. ~
1995 J L h r y Sci 781142-1153
1142
EFFECTS OF
1143
FISH MEAL ON MILK FAT
of PUFA in diets of lactating cows decreased the molar percentage of ruminal acetate and milk fat percentages, but noted that differences in ruminal VFA patterns were small compared with the large decrease in milk fat, suggesting that ruminal escape of PUFA may have allowed it to alter postruminal lipid metabolism. Infusion of cod liver oil into the rumen or abomasum also decreased milk fat percentage of lactating cows (22). These data support the postabsorptive effect of PUFA on lipid metabolism, as proposed by Vannan et al. (28), which includes decreased uptake of plasma fatty acids by the mammary gland. Effects of marine lipids and, more specifically, long-chain n-3 PUFA on lipid metabolism have been extensively researched. Eicosapentaenoic acid has been shown to inhibit activity of hepatic lipogenic enzymes (6,7). Our laboratory has found that cows fed FM had significantly higher plasma eicosapentaenoic acid and decosahexaenoic acid than These data suggest FM fatty cows not fed M. acids may alter post absorptive lipid metabolism. Therefore, a series of studies was conducted to reevaluate the use of FM as a protein source for lactating dairy cows. In all experi-
ments, a primary goal was to determine site of action by which fish meal has its negative effect on milk fat production. MATERIALS AND METHODS Ewrlmsnt 1
Eight Holstein cows were assigned to two concurrent replicates of a 4 x 4 Latin square, which included four cows per replicate receiving four different diets over four consecutive 20-d periods. Cows averaged 131 f 14 d of lactation and 27 f 2 kg of milk/d at the start of the study. Diets were T M R based on corn silage (Table 1) and were fed twice daily (O600 and 1500 h) in amounts adequate to ensure 10% om. Fish meal was substituted for corn gluten meal on an isonitrogenous basis. Cows were adjusted to diets for a 7-d transition period, followed by a 13-d collection period. Milk yield and DMI were recorded daily. Milk samples were collected at consecutive milkings on d 7 and 13 of the collection period and placed in sealed plastic bags containing potassium dichromate (150 mg per sample) as a preservative. Samples were stored at room
TABLE 1. Ingredient and chemical composition of diets.' Experiment 1 Composition
0% FM
2.6%
FM
5.2% FM
7.8%
FM
Basal diet Experiment 2l In situ Infusion C FM FO ,.- -- ----,
Ingredient Alfalfa silage Barley silage Corn silage thy corn High moisture corn Dried brewers grain soybean meal
(4%) Corn gluten meal FM Mineral mix Limestone chemical
DM, % ADF CP
... ...
... ...
... ...
...
62.3
62.9
62.0
61.9
...
...
35.8
...
...
36.1
77.9
37.5
...
37.6
8.2
51.9 16.6
...
...
...
16.4
...
35.8
... 37.3
...
...
...
20.3
20.5
20.2
20.2
...
...
14.9
15.0
14.9
...
...
...
...
...
3.6
...
...
...
10.1 7.3
10.3 3.7 2.6
10.1 2.5
...
12.2
7.8 2.9
7.5 3.7
...
7.5
7.5 3.9
...
... ...
42.0 25.2 17.2
5.2
...
10.1 7.8
...
...
...
1.7
.8
...
... ...
...
...
...
...
44.0 25.1 17.5
44.0 24.7 17.3
42.0 24.4 17.2
48.9 24.2 17.4
...
...
19.4 17.5
... ...
3.8
...
.6
...
44.3 24.5 15.7
44.0 24.2 16.3
...
. . . .6 44.3 24.2 15.5
IC = Control, FM = fish meal, and FO = fish oil, Journal of Dairy Science Vol. 78. No. 5, 1995
1144
SPAIN ET AL.
temperature for 24 h before being analyzed with a four-channel spectrophotometer (Multispec Mark I; Foss Food Technology, Eden Prairie, MN) for milk fat, protein, and lactose by Virginia DHIA. Approximately 150 ml of ruminal fluid were sampled by aspiration via esophogeal tube 2 to 3 h after the p.m. feeding on d 12 of the collection period. A 5-ml aliquot was placed in a plastic tube containing four drops of concentrated sulfuric acid for ammonia analysis. Another 10-ml aliquot was stored frozen (-2O'C) in a separate plastic tube for VFA analysis. Also, on d 12 of the collection period, 30 ml of blood were sampled by jugular puncture. Ten-milliliter subsamples were placed in plastic tubes containing 100 units of heparin each. All samples were stored on ice for transport to the laboratory. Blood samples were centrifuged at 3220 x g for 20 min at 5'C. Plasma was decanted and stored at -2O'C until analyzed. Plasma used for fatty acid analysis was stored at -8o'C. Ruminal fluid was stored at -2O'C until analyzed. Plasma urea was determined colorimetrically (4). Samples of diet components and TMR were collected weekly, and DM was determined by drying samples at 1WC in a forced-air oven for 24 h. Dried samples were ground to pass through a 2-mm screen (Wiley mill; Arthur H. Thomas, Philadelphia, PA). Kjeldahl N (1) and ADF (5) were measured on ground subsamples. Plasma lipids were extracted with ch1oroform:methanol (2: 1; volhol). Methyl esters of fatty acids from plasma, menhaden oil, duodenal ingesta, and milk fat were prepared by using 14% boron trifluoride and analyzed by capillary GLC (23). The methyl esters were extracted in hexane for chromatographic analysis by an HP 5890A gas chromatograph equipped with a flame ionization detector and integrator (Hewlett-Packard Co., Avondale, PA). A DM 225 (25% cyanopropylphenyl) fused silica capillary column, 30 m x .25 mm i.d. (J & W Scientific Co., Rancho Cordova, CA), was used with helium as the carrier gas. The initial oven temperature of 196'C was held for 12 min and increased at a rate of .9"U min until the final temperature of 214°C was reached. The total GLC run time was 40 min. An external standard mixture prepared from known amounts of triacylglycerols and methylated fatty acids (Nu Check-Prep, ElyJournal of Dairy Science Vol. 78, No. 5, 1995
sian, MN) was used to obtain retention times and to develop the calibration table. Heptadecaenoic acid (17: 1, voVvol) was added to the external standard mixture and to all samples, except the duodenal digesta, as the internal standard. Fatty acid values are presented as micrograms per milligram in the menhaden oil and milk fat and as micrograms per milliliter of plasma. Fatty acid values in duodenal digesta are reported as area percentages. Acidified ruminal fluid samples were thawed at room temperature, centrifuged at 3220 x g for 20 min, and analyzed for ammonia (13). Ruminal VFA concentrations were determined by the method previously used by Spain et al. (26). Data were analyzed statistically using the general linear models procedure of SAS (16). The statistical model used was Y = p + Ci + Trt, + Pdk + (TrtPd),k, where p is the population mean, C, is the cow i., Trt, is treatment j, Pdk is period k, and (TrtPd)jk. is the interaction between treatment j and penod k (10). Treatment means were tested by error means squares. Differences among means were evaluated by Tukey's procedure (10). Linear and quadratic response relationships were measured by orthogonal contrasts. In Situ Characterization of FM
Three ruminally fistulated cows were housed in a tie-stall barn and were individually fed a TMR (Table 1) twice daily. Approximately 10 g of FM were placed in dacron polyester bags (143 cm2) with a pore size of 57 pm. Bags were fastened to metal chains attached to nylon cords with three bags per incubation period per cow. Ruminal resident times for the bags were 2, 4, 8, 12, 24, 48, 72, and 120 h. At the beginning of incubation, bags were placed on the ventral floor of the rumen, and they were allowed to mix and be moved within the rumen. Bags were placed in the rumen in reverse order over time to allow the simultaneous removal of all bags. Upon removal, bags were individually rinsed to remove particles from the bag exterior. Bags were then placed in a continuous tap water rinse system for 24 h. During the continuous rinse, the system was emptied twice and refilled with fresh water. After rinsing, bags were suspended in a forced-air oven
1145
EFFECTS OF FISH MEAL ON MILK FAT
and dried for 48 h at 65°C. Dried residues were composited for each cow by residence time period. Composites were ground through a .5mm screen. Each composite was analyzed for DM and CP (1) as previously described. Lipid was analyzed by refluxing residue with petroleum ether for 1 h, and solvent was then removed (Soxtec System HT; Tecator, Herndon, VA). The remaining lipid was then weighed. Marlne Lipid infusion
Six lactating Holsteins (130 f 9 d of lactation) were assigned to two concurrent replicates of a 3 x 3 Latin square. The design included three 10-d periods during which cows received one of three treatments. Cows had been surgically fitted with ruminal fistulas and duodenal T-type cannulas. At the initiation of experiment, mean milk yield was 34.0 f 3.9 kg/d. Cows were housed in a tie-stall barn. Treatments were corn gluten meal administered into the rumen, corn gluten meal plus fish oil administered into the rumen, and corn gluten meal administered into the rumen plus fish oil infused into the duodenum. All cows were fed the same basal diet twice daily for 7 d prior to initiation of and throughout the 30-d experiment (Table 1). Cows were fed 30% of their daily ration at O600 h and the remaining 70% at 1500 h. Orts were removed daily and sampled for individual cows every 3 d of each period. The corn gluten meal was a mixture (86:14, d w t ) of corn gluten meal and limestone. The 341-g dose was placed into the rumen twice daily. The amount given was equivalent to the protein, calcium, and, when appropriate, fish oil in an equal amount of FM. Fish oil was a crude menhaden oil, and cows were dosed twice daily (48 d d ) 2 h after each feeding. Cows receiving the duodenal infusion of fish oil received one-half of the dose 2 h after feeding with the remainder given 1 h later. The fatty acid composition of the crude fish oil is shown in Table 2. Cows were milked twice daily (1200 and 2400 h), and yield was recorded. Milk samples were collected at consecutive a.m. and p.m. milkings on d 4.7, and 10 of each experimental period. Samples were handled and analyzed as described for Experiment 1. On d 10, 1 L of milk was collected at the p.m. milking and
TABLE 2. Fatty acid composition of crude Menhaden fish Oil'.
Fatty acid
Wmg)
c12:o c14:O
Trace 85 5 175 100 12 7 30
C18:l C18:l c18:2 c18:2 C18:3n6 C18:3n6 C18:3n3 C18:3n3 C Cm m 00
Cm1 Cm1
c20:2 c20:2
c20:3n6 c20:3n6 CD4n6 CD4n6 C20:3n3 C20:3n3 C205n3 C205n3 c22: c22: 1 1n9 n9 c22:6n3 C22.hn3
64 48 2 7 3 8 Trace 10 Trace Trace 112 Trace
59
IMenhaden fish oil was an unrefined, crude oil that was used to compare with residual crude oil associated with fish meal.
transported to the laboratory at ambient temperature. A 225-m1 aliquot was centrifuged at 9500 x g for 1 h at 10°C. The lower aqueous layer was aspirated by suction to isolate the fat layer. The milk fat was placed in individual plastic bags, which were sealed in larger plastic bags and stored at -7O'C until fatty acid analysis. Samples of ruminal fluid and blood were collected 2 h postdosing on d 10. Ruminal fluid was collected from various regions of the rumen and filtered through four layers of cheesecloth. Ruminal pH was measured immediately upon collection of the fluid. Ruminal fluid was mixed and subsampled for ammonia and VFA analysis, as outlined for Experiment 1. Blood samples were collected by puncture of the jugular vein, transferred to plastic tubes containing 200U of heparin, and processed as described for Experiment 1. Plasma was decanted and stored in plastic tubes at -20°C until analyzed. Plasma samples used for fatty acid analysis were stored at -70°C until analysis, as previously described. One liter of continuous duodenal digesta samples was collected at 0100, 0500, 0900, Journal of Dairy Science Vol. 78, No. 5, 1995
1146
SPAIN ET AL.
1300, 1700, and 2100 h over a 3-d period, representing 4-h intervals in a 24-h day. Duodenal digesta pH was measured electronically using a pH meter. Duodenal samples were stored in sealed plastic cups at -2O'C. For analysis, duodenal samples were thawed to room temperature and composited by cow for each period. Dry matter content was determined as the loss of weight following freezedrying for 120 h. Dried samples were ground to pass through a .5-mm screen. Acid detergent fiber was measured by the method of Goering and Van Soest (5). Acid-insoluble ash was determined by ashing the ADF residue at 500'C for 4 h. Acid-insoluble ash was a reference marker used to calculate postruminal flow. Fatty acids of duodenal DM were extracted by the method of Outen et al. (20) and analyzed by GLC. Duodenal cytosine was determined as described by Zerbini et al. (31). Diet component samples were collected weekly. The ADF and acid detergent-insoluble ash were measured as described for duodenal samples. Kjeldahl N was measured and reported as CP. The experimental model was Y = p + replicate + cow(replicate) + period + treatment, where p = treatments were tested by the error means square, and means were compared by Tukey's procedure (15). Statistical analysis was by the general linear models procedure of SAS
twice daily, and yield was recorded at each milking. Milk samples were collected at four consecutive milkings on d 16, 17, and 18 and analyzed for fat, protein, and lactose as described previously. Blood and ruminal samples were taken 2 h following the morning feeding and analyzed for plasma urea N, plasma long-chain fatty acids, ruminal ammonia, and ruminal VFA using procedures reported earlier. Dietary treatments were arranged in a 3 x 3 Latin square design with four replicates conducted concurrently. The experimental model was defined as Y = p + replicate + cow(rep1icate.) + period + treatment + residual with treatment means squares tested by residual means squares (10). Means were separated by Tukey's procedure (15). Statistical analysis was by the general linear models procedure of SAS
(W. RESULTS AND DISCUSSION Experiment 1
In Experiment 1, DMI was greatest when diets contained 2.6% FM and was lowest when 7.8% FM was included in the diet, resulting in a quadratic response (Table 3). Although inclusion of FM significantly affected DMI, the differences in intake were not of sufficient magnitude to alter milk production. Milk fat (W. percentage decreased linearly as intake of FM increased from 3.5% for controls to 3.0% for Experiment 2 cows fed the 7.8% FM diet. Milk protein and Twelve lactating Holstein cows (mean lacta- lactose percentages were not affected by the tion, 3.1) were assigned to one of four repli- diet. cates of a 3 x 3 Latin square design. Cows Ruminal N H 3 N appeared to respond in a were housed in a tie-stall barn and fed one of quadratic manner (P < .15); the value was three diets over three 18-d periods. Diets were highest (6.6 mgldl) when 2.6% FM was fed, T M R based on corn silage and alfalfa haylage and lowest (4.8 mg/dl) when 0 or 7.8% FM (Table 1). Fish oil was mixed with a corn was fed. Treatment means paralleled the patgluten meal and mineral premix, placed in tern for DMI (P < .15). Concentrations of plastic bags, and stored at 6'C. This premix plasma glucose (70 mgldl) and urea (21 mddl) contained corn gluten meal (80.0%), fish oil were also similar across the diets. (7.2%), and limestone (12.8%), making the Differences in milk fat cannot be attributed content of fish oil and calcium in the mix to altered ruminal fermentation. Total concensimilar to that in FM. Ethoxyquin (500 ppm) trations of VFA in ruminal fluid ranged from was added as an antioxidant during premixing 145.3 to 165.6 mM but did not differ with diet. of the fish oil with the corn gluten meal plus Individual VFA concentrations also were unlimestone. Cows were fed twice daily at O600 affected by diets. The molar percentage of and 1500 h in amounts to ensure 10% orts. butyrate was higher (P < .05; 13.5 vs. 12.5 Orts were recorded 4 dwk. Cows were milked moVl00 mol) for cows fed 5.2% FM than for Journal of Dairy Science Vol. 78, No. 5, 1995
1147
EFFECTS OF FISH MEAL ON MILK FAT TABLE 3. Effect of fish meal (FM) on DMI. milk yield, and milk composition in Experiment 1. Dietary treatments Item
0% FM
2.6% FM
5.2% FM
7.8% FM
SEM
DM1.l kg/d Milk, kg/d Fat,z % Protein, 96 Lactose. %
20.w 25.3 3.4
21.21 26.2 3.2ab 3.4
20.1b 25.5 3.1b 3.4
19Sb 25.6 3.P 3.3
.1 .4 .04 .01
4.1
4.1
47
47
3.53
apbMeans in same row followed by different superscripts differ (P < .05). lsignificant quadratic response to dietary treatment. ZSignificant linear response to dietary treatment.
cows fed 7.8% FM. The acetate to propionate ratio ranged from 2.6 to 2.8 but was not different among treatments. Jenny et al. (8) reported that decreased milk fat was associated with shifts in ruminal VFA patterns and concurrent increase in concentrations of plasma glucose. Plasma glucose was highest for cows fed high FM diet (73.6 mg/dl) but was not different from means of other treatments. Cows fed linear increments of FM exhibited a concurrent linear decrease in milk fat content in the absence of significant changes in milk volume, ruminal VFA patterns, or plasma glucose. Inclusion of FM in increasing concentrations did alter plasma PUFA profiles (Table 4).
TABLE 4. Effect of different amounts of fish meal increasing amounts of FM in Experiment 1.
Unsaturated fatty acids C16:ln7, C18:3n3, and C22:6n3 increased linearly (Pc .05) as FM intake increased. In contrast, C18:3n6 and C20:3n6 decreased linearly ( p < .05) as FM intake increased. Saturated fatty acids and the fatty acids C18:ln9 and C18:2n6 were not altered by the diet. It is important to note that C20:5n3 and C22:6n3 survived microbial biohydrogenation, thus allowing for the absorption of these long-chain PUFA. With regard to metabolic impact of these n-3 PUFA, reports (6, 7, 30)indicate that inclusion of n-3 PUFA in experimental diets of nonruminants resulted in a decreased activity of several key lipogenic enzymes. Activities of acetyl coenzyme A car-
C204n6, C20:5n3,
on plasma concentrationsof long-chain fatty acid of cows fed Diet
Fatty acid
0% FM
2.6% FM
5.2% FM
7.8% Fhl
SEM
237 19b 19 359 167 622
38 1 2 I89 18 21 1 1 1 3 2
~~
c16:O C16:1A c17:O C18 C18:ln9 C18:2n6 c18:3n6A C18:3n3A C203116~ CU):4n6A C205dA c224n6 C226n3A
239 12' 18 384 126 708 16 22a 436 38 91 5 9
otg/ml) 187 14l 15 292 117 713 11 27.b 35.b 47 30h 5 17
198
123 16 318 121 690 14 233 4o.b
41 19*b 5 13 ~~
10
3Ib 31b 51
42C 4 22
1
2
__
l.b~cMeansin same row followed by different letters differ (P < .05). ASignificant linear effects to diet (P < .Os).
Journal of Dairy Science Vol. 78, No. 5, 1995
1148
SPAIN ET AL.
TABLE 5. Disappearance1 of DM, CP. and ether extract of fish meal during in situ incubation in the rumen.
0 2 4 8 12 24 48 72 120
(%)
28.9 29.8 31.4 32.5 37.1 50.2 59.8 71.8
17.6 17.9 22.5 22.0 30.6 49.7 64.4 82.8
75.2 84.2 83.9 84.4 92.1 92.1 88.9 92.9
Percentage disappearance = 100 minus the percentage remaining in in situ bag at a given time.
ble 5). These results were similar to those reported by Zerbini et al. (31). In contrast to DM and CP, ether extract was removed very rapidly; 75% was removed after 2 h, and 92% of original lipid was removed after 24 h. Obviously, fat in FM is readily a part of the ruminal environment and may affect microbial metabolism in the rumen (17). These data do not address the availability of specific fatty acids, which may differ in their release and uptake. Individual fatty acid residues may not alter total flow to postruminal absorptive sites. The potential of the n-3 PUFA to alter lipid metabolism makes this lipid material especially important to evaluate. Firh 01 inturion Trial
The DMI averaged 19.7 kg/d and was not different among treatments (Table 6). Daily milk yields also were similar among treatments and averaged 31.6 kg/d. No differences occurred in milk composition and component yields among treatments. Ruminal VFA patterns were also similar across diets (Table 7). 2). Although total VFA concentration tended to be lower for ruminal infusion of fish oil, the in Situ Trial differences were not significant. Pennington Data from Experiment 1 suggested that marine PUFA passed from the rumen; therefore, and Davis (22) decreased milk fat content by the in situ experiment was conducted to evalu- infusing 225 g of destearinated cod liver oil ate ruminal degradation characteristics of men- into the rumen or abomasum. The change in haden FM. Preincubated Fh4 contained 94.3% milk composition occurred without changes in DM, 61.8% CP, and 9.4% fat, which were ruminal VFA patterns. Varman et al. (28) similar to previously reported values (9). By 24 reported similar results from dietary fish oil h, 37 and 30.6% of the original DM and CP, supplement fed to lactating cows. Ruminal respectively, were removed from the bag (Ta- acetate decreased slightly by inclusion of fish boxylase and fatty acid synthetase were greatly reduced for mice fed diets high in eicosapentaenoic acid (C205n3). This fatty acid is also found in plasma of mice fed marine oil containing high amounts of C205n3 (6). Up to 25% of the fatty acids in FM are n-3 PUFA (Table
TABLE 6. Effect of ruminal or duodenal infusion of fish oil on intake, milk yield, composition, and component yields. Infusion method Item
None
Ruminal
DMI Milk Fat Protein Lactose
19.7 31.3
20.2 31.5 1 .o 1 .o 1.5
Duodenal
SEM
19.2 32.0 1 .o 1 .o 1.5
.2 .2
3.2 3.1
3.1 3.1
.04 .01
4.8
4.1
.01
Wd)
I .o
.o
1 1.5
.o 1 .01 .o1
(W Fat Protein Lactose
3.2 3.1 4.7
Journal of Dairy Science Vol. 78, No. 5, 1995
1149
EFFECTS OF FISH MEAL ON MILK FAT
TABLE 7. Effect of site of infusion of fish oil in the rumen or duodenum on ruminal VFA concentrations, ratios, and pH. lnfusion method Item
None
Ruminal
Total Acetate (A) Propionate (P) Isobutyrate Butyrate Isovalerate Valerate A:P Ruminal pH
150.9 82.0 40.2 1.6 20.9 3.1 3.2 2.2 5.9
140.1 80.0 33.5 1.6 18.7 3.1 3.4 2.5 5.8
Duodenal
SEM
150.7 86.8 34.7 1.7 20.5 3.5 3.6 2.7 6.0
2.1 1.6 2.0 .03 .1 .1 .1 .2 .02
(mM)
oil but not of the magnitude associated with a large decrease in milk fat content that they observed. V m a n et al. (28) reported that arteriovenous differences in mammary triglyceride uptake decreased, suggesting a possible postabsorptive effect of fish oil fatty acids. In contrast, Storry et al. (27) reported decreased ruminal acetate in cows fed 300 g of unprotected cod liver oil, which resulted in decreased milk fat production. In all of these reports, fish oil amounts were greater than those fed in this experiment (-50 gld) and in Experiment 1 (-150 g/d). In Experiment 1, corn silage was the only forage, but alfalfa provided 16 and 35% of the DM in the infusion experiment and Experiment 2, which may have resulted in less milk fat response, as observed by Broderick (3). Duodenal digesta flows (14.5 kgld) were not different among treatments. Duodenal digesta pH and cytosine concentration were also unaffected by treatment. These results, in addition to the lack of differences in DMI and milk yield, further support the conclusion that fish oil did not alter ruminal fermentation in the present study. Although small differences were measured in fatty acid contents of duodenal digesta, these differences would be expected to have little metabolic consequence for the animal (Table 8); however, infusion of fish oil altered plasma fatty acid concentrations, and the response differed among treatments (Table 9). Saturated fatty acids and the monoenoic Cl8:ln9 were not different because of treatment. Palmitoleic acid (C16:ln7) was increased
(P-c .09) by duodenal infusion of fish oil. This small difference was probably of little consequence to the cow; however, PUFA were markedly changed by infusion of fish oil. yLinolenic (cl8:3n6) was decreased by infusion of fish oil compared with that of control cows. The COnCen@i3tiOnSOf C204n6, C20:5n3. and c 2 2 6 f l were all increased by duodenal infusion of fish oil. Apparently, because of microbial metabolism in the rumen, administration of fish oil into the rumen failed to cause changes in plasma fatty acid profiles. These differences associated with the source of oil between Experiment 1 and the infusion study suggest differences in ruminal stability or microbial metabolism of fat in fish oil versus FM. Although plasma fatty acids were altered, milk fatty acid composition did not differ from C 4 to c l 8 including C18:Ov C 1 8 : l ~ c 1 8 : 2 ~ C18:3n6, and C18:3n3. Only trace amounts above c 1 8 were found. Studies in which whole oil seeds or protected lipids have been fed have shown significantly altered fatty acid composition of milk (13, 16). Pennington and Davis (22) increased unsaturated fatty acid composition of milk with infusion of fish oil. Nicholson and Sutton (18) reported a linear increase of c16:1 in milk fat as fish oil increased. Pennington and Davis (22) also reported increases in the presence of unidentified 20-carbon fatty acids after cows were infused with fish oil. Dosage in the present study was lower than those previously reported but was calculated to reflect normal lipid intake from a diet supplemented with FM. This lack of reJournal of Dairy Science Vol. 78. No. 5, 1995
1150
SPAIN ET AL.
TABLE 8. Effect of site of infusion of tish oil on fatty acid composition of duodenal digesta. Infusion method Fatty acid c4:0 c6:0 C8:O
c1o:o c120 c13:0 c140 c141
c15:o C1S:l c16:0
c16:I c17:0
Cl8:O Cl8:l Cl8:2 I8:3n6 C18:3n3
c20:o
None .8* .2 .4
.2 .4 .1 1.5 .2 1.o .5 18.3 .3 1.1 51.8 10.8 7.3
Trace .7 .8a
Ruminal
Duodenal
.4b .2 .4 .2 .4 .2 I .7 .2 1.o .4
18.9 .3 1.2 48.7 14.4 7.7 TraCe .E 1.Ob
SEM
.5* .2
.05 .01
.4
.02
.2 .4 .2 1.4 .2 1.o .5 17.8 .2 1.1 45.5 11.3 10.4 TraCe
.02 .01 .02 .04
.8 .Ea
.02
.01 .05 .2 .03 .02 1.o 1.o .5
.02 .02
*.bMeans in same row with different superscripts differ (P < .05).
sponse, compared with previously reported studies (14), may also result from the preferential use of saturated and monoenoic fatty acids for milk triglyceride synthesis; furthermore, length of experimental periods might have been inadequate to allow equilibration of fatty acids and maximum metabolic responses to changes in plasma fatty acid concentrations.
Neither ruminal total VFA concentrations (96.4 mM) nor individual VFA differed with diet. Acetate:propionate ratios averaged 3.4.
This lack of response from the inclusion of FM or fish oil agrees with published results (22). Nicholson and Sutton (18) have reported that the addition of cod liver oil to diets of dauy cows reduced ruminal acetate and butyrate concentrations; however, fiber influenced VFA concentrations, and the change was more proExperiment 2 nounced for the low roughage diets than for With the apparent differences in digestion medium or high roughage diets. Fiber source between FM and fish oil, this experiment was may also influence response to M or fish oil. conducted to compare feeding free oil or oil Broderick (3) reported FM addition to diets indigenous in FM. Diet did not affect DMI or based on alfalfa silage did not change milk milk yield (Table lo), which agrees with previ- composition. Alfalfa supplied 36% of the DM ous experiments showing no differences on in Experiment 2. Differences because of yield when FM was substituted for corn gluten forages may reflect differences in ruminal fermeal. Milk fat decreased .1 percentage units by mentation. Differences in microbial fermentaFM and fish oil, but total fat yield was equal. tion in the rumen, caused by soluble and strucNo other differences in component concentra- tural carbohydrates, may cause hormonal tions or yields were measured. When Oldham changes in the animal and alter fatty acid et al. (19) substituted FM for urea in diets of metabolism (8), which may change the effect lactating dairy cows, milk protein yields were of n-3 fatty acids on lipid metabolism. significantly increased. Substitution of FM for Shifts in plasma fatty acid concentrations soybean oil meal did not alter milk composi- were similar to those in the previous two tion. studies (Table 11). The FM tended to lower Journal of Dairy Science Vol. 78, No. 5, 1995
1151
EFFECTS OF FISH MEAL ON MILK FAT TABLE 9. Effect of site of fish oil infusion on long-chain fatty acid composition in plasma. Infusion Fatty acid
None
C160
28 1 14 24 443 181 1loo 301 66 69a 17a 8 20 Trace.
Ruminal
Duodenal
SEM
OCW) C16I c17:0 c18:0 c18:1 c18:2 C18:3n6 C20:3n6 C204n6 C205n3 C22:4n6 C22:5n3 C22:6n3
258 15 24
294 24 25
409
440
193 1163 25b 69 701 2ga 9 19 2'
174 1126 22b 65 82b 64b
a*bMeans in same row with different superscripts differ
C18:2n6$ C18:3n6. and C20:3n6. In contrast, C16:1, Cl8:3n3, C20:4n6*.C20:5n39 and C22:6n3 were in-
creased by the inclusion of FM in the diet. Fish oil increased only C20:5n3 when compared with fatty acids of the control diet. These responses agree with the previous work done at our lab with a similar percentage of FM. These results indicate a difference exists in the ruminal characteristic between the FM and fish oil as sources of supplemental marine lipids; however, in previous experiments, FM addition to diets based on corn silage was accompanied by decreased milk fat content (26). In the present study, milk fat decreased only slightly with a similar increase in plasma n-3
20 2 2 30 11 18 1 2 2 1
10
1
28 12b
2 1
(P c .05).
fatty acids. Potential for forage source to influence yield responses should be considered (3). CONCLUSIONS
Fish meal altered milk fat percentage when fed in diets based on corn silage. This change in milk composition was not associated with changes in patterns of ruminal VFA. Increased concentrations of n-3 fatty acids were measured with the inclusion of FM, suggesting a protection of these PUFA from microbial metabolism in the rumen. Use of fish oil as a comparison for FM seemed to produce a
TABLE 10. Effect of dietary treatment on milk yield, DMI, milk composition, and component yields of cows on Experiment 2. Diet
Ikm
Control
Fish meal
Fish oil
SEM
Milk, kg/d
30.1 21.6
30.3 21.2
30.1 21.4
.2 .3
3.8 1.1
3.7 1.1
3.7 1.1
.03 .01
3.1 .9
3. I .9
3.1 .9
.01
4.8 1.4
4.8 1.4
4.8 1.4
.01
DM, kg/d Milk fat 5%
kdd Milk protein 5%
Wd Milk lactose 5%
kg/d
.01
.01
Journal of Dairy Science Vol. 78, No. 5, 1995
1152
SPAIN ET AL
TABLE 11. Effect of dietary fish meal or fish oil fed to lactating cows on fatty acid concentrations in plasma of cows in Experiment 2. Diet
Item
Control
c16:0 c16:1 c17:0
268 19. 23 397 172 1331 31 142 74 7 9 26' 9 22 4'
Fish meal
Fish oil
SEM
286 26b 24 391 182 1195 23 167 65 87b 60b 12 27 18b
8 .7 .7 12
Wm)
C18:O
C18:l C18:2 C18:3n6 18:3n3 C20:3n6 C20:4n6 C205n3 C22:4n6 C225n3
C226n3
288 2or 24 419 175 1334 29 146 74 723 32b 11 22 4'
5
42 1 4 2 2 2 .7 1 1
aJ'Means in same row followed by different a superscript differ (P < .05).
different response based on changes in plasma fatty acid profiles. The differences in the plasma fatty acid response between ruminal infusion or dietary fish oil, compared with duodenal infusion of fish and dietary FM,support this observation. The exact role of n-3 PUFA on milk lipid yield should be investigated further. At sufficiently high amounts of dietary fatty acids from fish, ruminal activity would probably be altered, as has been observed with other unsaturated fatty acids. However, VFA patterns were not altered in this study, even though milk fat declined. Fish oil is released readily from FM.Perhaps amounts infused or fed as FM were too small to elicit a response, even though the amounts fed coincided with practical feeding amounts. The inclusion of alfalfa in the diet may also have influenced the response. The ability to manipulate milk fat yield without excessive grain feeding would be attractive, given current changes in milk fat demand. The interaction of forage base and response to n-3 PUFA still needs investigation. REFERENCES 1 Association of Official Analytical Chemists. 1980. Official Methods of Analysis. 13th ed. AOAC, Washington, DC. 2Atwal. A. S., and J. D. M e . 1992. Effects of fish mal feeding to cows on digestibility, milk production, and milk composition. J. Dairy Sci. 75502.
Journal of Dairy Science Vol. 78, No. 5 , 1995
3Broderick. G. A. 1992. Relative value of fish meal versus solvent soybean meal for lactating dairy cows fed alfalfa silage as sole forage. J. Dairy Sci. 75:174. 4 Coulombe, J. J., and L. Favreau. 1963. A new simple semimicro method for calorimetric determination of urea. clin. Chcm. 9:102. SGoeMg, H. K., and P. J. Van Soest. 1970. Forage Fiber Analyses (Apparatus, Reagents, Procedures, and some Applications). Agric. Handbook No. 379. ARSUSDA, Washington, DC. 6Huzberg. G. R.. and M. Rogerson. 1988. Hepatic fatty acid synthesis and triglyceride secretion in rats fed fructose or glucose-based diets containing corn oil, tallow or marine oil. J. Nutr. 118:1061. 7 Ifitani, N., K.Inoguchi, M. Endo, E. Fudude, and M. Morita. 1980. Identification of shellfish fatty acids and their effects on lipogenic enzymes. Biochem. Biophys. Acta 68:378. 8 J e ~ y B. . F., C. E. Polan, and F. W. Thye. 1975. Effects of high grain feeding and stage of lactation on Serum insulin, glucose, and milk fat percentage in lactating cows. J. Nutr. 104:379. 9 Klopfenstein, T. 1991. W i n g animal protein products. Page 60 in Pmc. Alternative Feeds for Dairy and Beef Cattle Symp., St. Louis, MO. E. R. Jordon. ed. Sponsored by USDA-Extension Service, industry support, and Univ. Missouri Ext., Columbia. IOLcntner, M., and T. Bishop. 1986. Experimental Design and Analysis. Valley Book Co., Blacksburg, VA. 11 Mbtysaari, P. E.,C. J. Sniffen, T. V. Muscato. J. M. Lynch, and D. M. Barbano. 1989. Performance of cows in early lactation fed isonitrogenous diets containing soybean meal or animal by-product meals. J. Dairy Sci. 72:2958. 12 Mattos, W., and D. L. Palmquist. 1974. Increased polyunsaturated fatty acid yields in milk of cows fed protected fats. J. Dairy Sci. 57:1050.
EFFECTS OF FISH MEAL ON MILK FAT 13 McCullough, H. 1967. The determination of ammonia in whole blood by direct colorimetric method. Clin. Chem. Acta 17:297. 14 Moore, J. H., and W.W.Christie. 1980. Lipid metabolism in the mammary gland of ruminants. Rog. Lipids Res. 17:347. 15 Mosteller, F.. and J. W. Tukey. 1977. Data Analysis and Regression. Addison-Wiley, New York, NY. 16 Murphy, J. J., and D. J. Morgan. 1978. Effect of inclusion of protected and unprotected tallow in dairy rations on cow performance. A n i . Prod. 32:368. 17National Research Council. 1989. Nutrient Requirements of Dairy Cattle. 6th rev. d.Natl. Acad. Sci., Washington, DC. 18Nicholson. J.W.G..and J. D. Sutton. 1971. Some effects of unsaturated oils given to dairy cows with rations of different roughage to concentrate. J. Dairy Res. 38:363. 19Oldham. J. D., D. J. Napper, T. Smith, and R. J. Fulford. 1985. Performance of dairy cows offered isonitrogenous diets containing urea or fish meal in early and mid-lactation. Br. J. Nutr. 53:337. 20Outen. G. E.,D. E. Beever. and 1. S. Fenlon. 1976. Direct methylation of long-chain fatty acids in feeds, digesta, and faeces without prior extraction. J. Sci. Food Agric. 27:419. 21 Palmquist. D. L., and T. C. Jenkii. 1980. Fats in lactation rations: review. J. Dairy Sci. 63:l. 22 Pennington, J. A., and C. L. Davis. 1975. Effects of intraruminal and intra-abomasaladditions of cod-liver on milk fat production in the cow. J. Dairy Sci. 58:49. 23 Phetteplace, H. W., and B. A. Watkins. 1990. Lipid measurements in chickens fed different combinations
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of chick fat and menhaden oil. J. Agric. Food Chem. 38:1848. 24 SAS@ User’s Guide: Statistics, Version 5 Edition. 1985. SAS Inst., Inc., C q , NC. 25Schwab. C. G.. C. K. Bo&. N. L. Whitehouse. and M.M.A. Mesbah. 1993. Amino acid limitation and flow to the duodenum at four stages of lactation. 1. Sequence of lysine and methionine limitation. J. Drury Sci. 75:3486. 26Spain, J. N., M. D. Alvarado, C. E. Polan. C. N. Miller, and M. L. McGilliard. 1990. Effect of protein source and energy on milk composition in midlactation dairy cows. J. Dairy Sci. 73:445. 27 Stony, J. E., P. E. Brumby. A. J. Hall, and B. Tuckley. 1974. Effects of free and protected forms of codliver oil on milk fat secretion in the dairy cow. J. Dairy Sci. 57:1046. 28 Varman, P. N., L. H. Schultz, and R. E. Nichols. 1968. Effect of unsaturated oils on rumen fermentation, blood components, and milk composition. J. Dairy Sci. 51:1956. 29 Wohlt, J. E., S. L. Chmiel. P. K.Zajac, L. Backer. D. B. Blethen, and J. L. Evans. 1991. Dry matter intake, milk yield and composition, and nitrogen use in Holstein cows fed soybean, fish. or corn gluten meals. J. Dauy Sci. 74:1609. 30Yang. Y.T., and M. A. Williams, 1978. Comparison of CIS-. Cm-, C 2 + ~ t u m t e dfatty acids in reducing fatty acid synthesis in isolated rat hepatocytes. Biochim. Biophys. Acta 531:133. 31 Zerbini, E., C. E. Polan, and J. H. Herbein. 1988. Effect of dietary soybean meal and fish meal on protein digesta flow in Holstein cows during early and midlactation. J. Dairy Sci. 71:1248.
Journal of Dairy Science Vol. 78, No. 5, 1995