Search
Author
Title
Vol.
Issue
Year
1st Page

Journal of Animal Science - Animal Nutrition

Ileal flows and apparent ileal digestibility of fatty acids in growing gilts fed flaxseed containing diets1

 

This article in JAS

  1. Vol. 91 No. 6, p. 2729-2739
     
    Received: Aug 23, 2012
    Accepted: Feb 22, 2013
    Published: November 25, 2014


    2 Corresponding author(s): cdelange@uoguelph.ca
 View
 Download
 Share

doi:10.2527/jas.2012-5783
  1. H. R. Martínez-Ramírez*,
  2. J. K. G. Kramer and
  3. C. F. M. de Lange 2
  1. Centre for Nutrition Modelling, Department of Animal and Poultry Science, University of Guelph, Guelph, ON N1G 2W1, Canada
    Agriculture and Agri-Food Canada, Guelph, ON N1G 5C9, Canada

Abstract

An experiment was conducted to quantify the ileal flow and apparent ileal digestibility (AID) of fatty acids (FA) in growing gilts fed corn, wheat, and soybean meal based diets without (CON) or with ground flaxseed (FS). A total of 20 healthy purebred Yorkshire female pigs, weighing approximately 25 kg BW, were allotted to 1 of 3 feeding regimens: R1 (n = 5 pigs), feeding a diet containing 10% FS between 25 and 50 kg BW and CON diet thereafter, R2 (n = 10 pigs), feeding CON diet between 25 and 85 kg BW and a diet containing 6% FS thereafter, and R3 (n = 5 pigs), feeding CON diet between 25 and 110 kg BW. Titanium dioxide was used as an indigestible marker to assess AID and ileal flows of crude fat and FA. At 110 kg BW, pigs were slaughtered and representative digesta samples were obtained from the distal ileum. Ileal flows and AID of crude fat and individual FA did not differ (P > 0.10) between R1 and R3, and therefore, results from these 2 feeding regimens were combined to give 2 dietary treatments (CON and FS). There were no treatment effects on AID of crude fat and the sum of all FA, SFA, or MUFA. However, the AID of individual SFA decreased with chain length (linear; P < 0.05) for both FS and CON. The AID of myristic acid (14:0), individual trans-18:1 FA (6t-8t-18:1 to 12t-18:1), myristoleic acid (9c-14:1), and palmitoleic acid (9c-16:1) were greater for CON than FS (P < 0.05) whereas no diet effect was observed for the AID of linoleic acid (18:2n-6; 80.2 and 86.1% for FS and CON, respectively) and α-linolenic acid (18:3n-3; 86.7 and 89.8% for FS and CON, respectively). Ileal flows of rumenic acid (9c11t-CLA), n-3 PUFA, and highly unsaturated FA (HUFA; arachidonic, eicosatrienoic, eicosapentaenoic, docosapentaenoic, and docosahexaenoic acids) exceeded their intakes, indicating net appearance of these FA in the upper gut of the pig. It remains to be determined whether enteric microbiota can elongate and desaturate 18:2n-6 and 18:3n-3 and isomerize 18:2n-6. The contribution of endogenous FA losses from the host to the ileal flow of these FA should also be considered. Further studies are needed to quantify production of CLA isomers and PUFA in the small intestine of pigs, specifically the n-3 HUFA, and to assess their contribution to the FA supply of the host.



INTRODUCTION

Fats from animal and plant origin can be used as dietary ingredients in animal production, providing means to increase dietary energy density (NRC, 1998) and manipulate fatty acid (FA) profiles of animal products (Wood et al., 2003). The diverse chemical structures of dietary fat as well as potential interactions with other dietary constituents and enteric microbiota can affect digestibility and use of FA in animals and humans (Jørgensen et al., 2000). In pigs, fat digestion and FA absorption take place before the distal ileum, and high rates of incomplete biohydrogenation of FA occur primarily in the lower gut (Duran-Montgé et al., 2007). Because of the relatively large content of SFA in animal fats (mainly palmitic acid and stearic acid), reduced digestibility has been observed for animal fats than plant oils that are generally rich in highly digestible MUFA (i.e., oleic acid, 9c-18:1) and PUFA (i.e., linoleic acid, 18:2n-6, and α-linolenic acid, 18:3n-3; Cera et al., 1988, 1989). Interactions of dietary fat with dietary fiber and minerals have also been shown to alter FA digestibility (Jørgensen et al., 1992; Dove, 1993; Li and Sauer, 1994).

Little attention has been given to ileal flows and apparent ileal digestibility (AID) of FA in pigs and biosynthesis of CLA isomers and metabolism of n-3 PUFA and n-6 PUFA in the upper gut of pigs. The fate of n-3 PUFA is highly relevant when feeding ground flaxseed (FS) to pigs for generating n-3 PUFA enriched pork (Romans et al., 1995a,b). The aim of this companion study was to characterize ileal flows and AID of FA in growing pigs fed a corn, wheat, and soybean meal based diet that included ground FS. A FS-free diet was used as a control (CON) and observations were made at 110 kg BW using an established method to analyze the ileal digesta content sampled at slaughter (Jeaurond et al., 2008).


MATERIAL AND METHODS

The University of Guelph Animal Care Committee approved the experimental protocol.

Animals, Diets, and General Design

A total of 20 healthy purebred Yorkshire gilts from 12 different litters, weighing approximately 20 kg BW were obtained from the Arkell Swine Research Station (Guelph, ON) and transported to the animal metabolism unit at the University of Guelph in blocks of 6, 8, and 6 pigs. The pigs were previously fed a pig starter diet (Table 1) ad libitum. General management of the individually housed pigs was described previously (Martínez-Ramírez et al., 2008).


View Full Table | Close Full ViewTable 1.

Ingredient composition and nutrient content of experimental diets1

 
Item Adaptation Grower flaxseed Grower control Finisher 1 control Finisher 2 flaxseed Finisher 2 control
Ingredient composition, % (as-fed)
    Corn 39.90 34.55 40.40 52.20 51.98 57.73
    Wheat 20.00 20.00 20.00 20.00 20.00 20.00
    Soybean meal 35.00 31.50 33.5 22.00 18.00 18.00
    Ground flaxseed 10.00 7.00
    Fat2 1.00
    Beef tallow 2.00 2.00 1.00
    l-Lys HCl 0.20 0.20 0.20 0.20 0.13 0.17
    dl-Met 0.12 0.08 0.08 0.04 0.03
    l-Thr 0.13 0.10 0.10 0.07 0.01 0.04
    l-Trp 0.02
    Lincomycin 443 0.10 0.10 0.10 0.10 0.10 0.10
    Limestone 1.15 1.15 1.12 1.12 1.06 1.06
    Dicalcium phosphate 1.30 1.20 1.38 1.15 0.60 0.73
    Salt 0.40 0.40 0.40 0.40 0.40 0.40
    Vitamin and mineral mix4 0.60 0.60 0.60 0.60 0.60 0.60
    Titanium dioxide 0.10 0.10 0.10 0.10 0.10 0.10
    Vitamin E5 0.02 0.02 0.02 0.02 0.02
Calculated nutrient content6
    DE, MJ/kg 14.6 14.6 14.7 14.7 14.5 14.5
    CP, % 22.7 22.7 21.9 17.5 16.9 16.0
    Crude fat, % 2.93 6.11 4.58 5.14 4.98 4.71
    Total Lys, % 1.40 1.32 1.32 1.00 0.88 0.87
    Digestible Lys,4 % 1.21 1.17 1.17 0.89 0.77 0.77
    Digestible Met + Cys, % 0.76 0.73 0.71 0.56 0.52 0.52
    Digestible Thr, % 0.85 0.82 0.81 0.62 0.53 0.54
    Digestible Trp, % 0.24 0.24 0.23 0.17 0.17 0.17
    Ca, % 0.78 0.78 0.77 0.70 0.60 0.60
    P, % 0.70 0.70 0.70 0.61 0.51 0.51
    Na, % 0.18 0.20 0.18 0.18 0.18 0.18
Analyzed nutrient content, %
    DM 92.8 89.8 90.5 88.7 88.4 90.2
    CP 20.2 23.1 21.3 17.9 17.2 16.9
    Ether extract (HCl hydrolysis) 4.67 7.36 5.32 5.92 6.32 4.81
    Ash 5.76 5.83 5.66 4.69 4.25 3.78
    Crude fiber 2.41 3.35 2.74 2.31 3.21 2.65
    Ca 0.82 0.81 0.87 0.65 0.59 0.49
    Ph 0.66 0.72 0.69 0.60 0.49 0.45
    Na 0.19 0.20 0.17 0.20 0.18 0.16
    K 0.73 1.03 0.95 0.75 0.73 0.72
1Diets were fed over these BW ranges: adaptation, up to 25 kg; grower, 25 to 50 kg; finisher 1, 50 to 85 kg; and finisher 2, 85 kg to 110 kg.
2Animal/vegetable fat blend.
3Supplied 44 mg of lincomycin/kg of diet as lincomycin-HCl (Pfizer Canada Inc., Kirkland, QC, Canada).
4Provided the following amounts of vitamins and trace minerals per kilogram of diet: vitamin A, 12,000 IU as retinyl acetate; vitamin D3, 1,200 IU as cholecalciferol; vitamin E, 48 IU as DL-α-tocopherol acetate; vitamin K, 3 mg as menadione; pantothenic acid, 18 mg; riboflavin, 6 mg; choline, 600 mg; folic acid, 2.4 mg; niacin, 30 mg; thiamine, 18 mg; pyridoxine, 1.8 mg; vitamin B12, 0.03 mg; biotin, 0.24 mg; Cu, 18 mg as CuSO4.5H2O; Fe, 120 mg as FeSO4; Mn, 24 mg as MnSO4; Zn, 126 mg as ZnO; Se, 0.36 mg as Na2SeO3; I, 0.6 mg as KI (DSM Nutritional Products Canada Inc., Ayr, ON, Canada).
5Concentratation of vitamin E: 500 IU/g of product
6Based on nutrient composition of feed ingredients according to NRC (1998).

Throughout the experiment, pigs (initial BW, 25.6 ± 1.9 kg) were fed 3 times a day (0700, 1500, and 2300 h), and feed intake levels were fixed at 95% of voluntary daily DE intake according to NRC (1998) and adjusted weekly based on BW. Within blocks, littermates were randomly assigned to different feeding regimens, and mean initial BW were balanced across feeding regimens. There were 3 feeding regimens: R1 (n = 5 pigs) consisted of a corn–wheat–soybean meal based diet containing 10% ground FS that was fed between 25 and 50 kg BW and a corn–wheat–soybean diet without FS (CON) diets thereafter until 110 kg BW, R2 (n = 10 pigs) consisted of feeding CON diet between 25 and 85 kg BW with 6% FS thereafter until 110 kg BW, and R3 (n = 5 pigs) consisted of feeding CON diets between 25 and 110 kg BW. The diet containing FS for R2 (6% FS) was prepared by blending the CON diet and the diet containing 7% FS in a 10:90 ratio. The inclusion of FS (6%) in the finishing diets was reduced relative to that in the grower diets to achieve similar cumulative intake of FS for R1 and R2 between 25 and 110 kg BW for the evaluation of postabsorptive use of FA.

The diets were formulated to ensure that intakes of AA, vitamins, minerals, and essential FA exceeded requirements according to NRC (1998; Table 1) and pelleted. Nutrient composition of the experimental diets was calculated based on feed ingredient compositions according to NRC (1998). Crystalline Lys, Met, Thr, and Trp were used to improve the dietary AA balance. Diets were formulated to have similar digestible Lys to DE ratios within BW ranges. Titanium dioxide was added to the experimental diets as an indigestible marker to assess the ileal flows and determine AID of crude fat and FA. Pigs were slaughtered at 110 kg BW. Details of the slaughter procedures were previously described (Martínez-Ramírez et al., 2008). Pigs were fed at 2300 h, fasted overnight, and killed at 0800 h the next day. Representative digesta samples were quickly taken from the last meter of the distal ileum by gently squeezing the isolated gut segment, and the samples were stored frozen at –80°C until they were analyzed (Jeaurond et al., 2008).

Sample Preparation and Chemical Analyses

Representative feed, ground FS, and digesta were sampled for subsequent nutrient analyses and stored frozen at –20°C. For FA analyses, representative feed samples were taken both at the start and the end of the experiment. For ground FS, samples were taken at the beginning of the experiment. Feed, FS, and freeze-dried digesta samples were analyzed for contents of fat, ash, DM, and titanium dioxide contents according to AOAC International (AOAC, 1997). Nitrogen contents of the experimental diets were determined using an induction furnace and a thermal conductivity nitrogen gas analyzer (Leco FP-528; Leco Corporation, St. Joseph, MI; AOAC 1997).

Fatty Acid Analyses

Preparation and FA analyses of feed, FS, and digesta samples was conducted according to Cruz-Hernandez et al. (2004) with slightly modifications. Briefly, neutral and polar lipids were extracted from approximately 10 g of homogenized and pulverized feed and FS samples by adding 100 mL of chloroform/methanol (1:1, vol/vol) and mixing it for 2 min using a mechanical mixer (Virtris-45 Homogenizer; Virtris, Gardiner, NY). The solution was then filtered and the organic solvents were removed from the extract using a rotary evaporator (Model RE-131; Büchi Laboratoriums-Technik, Flavil, Switzerland; Cruz-Hernandez et al., 2006). After most of the solvent was removed, 10 drops of benzene were added to the flask and the solvents were again removed using the rotary evaporator to remove the last traces of water by azeotropic distillation (Cruz-Hernandez et al., 2006). The total lipid content was determined gravimetrically and processed samples were then stored in chloroform at –80°C.

Freeze-dried digesta samples were ground under liquid N. Duplicate representative digesta samples (0.5 g) were placed in 15 mL culture tubes with screw caps and Teflon liners. After addition of the internal standard (pentadecanoic acid, 15:0) and 3 drops of benzene to dissolve the lipids, the samples were treated separately using an acid- or base-catalyzed hydrolysis and methylation procedure (Cruz-Hernandez et al., 2006). This duplicate procedure was done to ensure completeness of hydrolyzing the soaps and methylating all lipids using HCl as catalyst. Alkaline digestion followed by neutralization with HCl and methylation using trimethylsilyl diazomethane was performed to retain any acid labile CLA lipids in these mixtures.

Acid-Catalyzed Hydrolysis and Methylation.

Acid fat hydrolysis and FA methylation were performed by adding 5 mL of a 5% solution of HCl gas in anhydrous methanol (wt/vol) to freeze-dried digesta samples containing about 10 mg fat in a 15 mL test tube with cap and Teflon liners. The sample was then mixed by a vortex and heated for 1 h at 80°C (Cruz-Hernandez et al., 2004). After cooling to room temperature, 1 mL of water was added and the fatty acid methyl esters (FAME) were extracted 2 times using 1.5 mL of hexane. To clarify the layers, the samples were centrifuged using a clinical centrifuge (2,000 × g for 5 min at 4°C), and the upper layer was transferred to a new vial.

Saponification and Base-Catalyzed Methylation.

The base-catalyzed methylation procedure was performed by adding 4 mL of 0.5 N NaOH in anhydrous methanol to freeze-dried digesta samples containing about 10 mg of fat in a 15 mL test tube. Samples were then mixed by using a vortex and hydrolyzed for either 15 min (diets) or 30 min (digesta) at 80°C. After cooling to room temperature, approximately 4 mL of 2.5 M aqueous HCl was added to the solution to achieve a pH between 5.5 and 6.0. The FFA were extracted 2 times using 1.5 mL of hexane. To clarify the layers, the samples were centrifuged using a clinical centrifuge (2,000 × g for 5 min at 4°C). The hexane layers were combined and a portion of the hexane solution representing about 2 mg of FFA was placed into a new test tube. After removal of the hexane, 1 mL of benzene/methanol (4:1, vol/vol) was added and mixed. Dropwise addition of trimethylsilyl-diazomethane (TCI America, Portland, OR) was performed in the fume hood until a yellow color persisted. The tube was intermittently shaken gently for 15 min. Glacial acetic acid was then added by drops until the yellow color disappeared and the derivatizing agent was inactivated. After addition of 1 mL water, the FAME were extracted 2 times using 1.5 mL of hexane, and the combined hexane layers were transferred to a new vial.

Approximately 2 mg of the FAME produced by the base- and acid-catalyzed hydrolysis-methylation procedure were both purified by TLC using silica gel G plates (Fisher Scientific; Ottawa, ON, Canada) and developed using the solvent mixture hexane/diethyl ether/acetic acid (85:15:1 by volume). The FAME band was removed from the TLC plate after visualization by spraying the plate with a 2’,7’-dichlorofluorescein solution (0.001% in methanol). No triacylglycerols, cholesterol esters, FFA, or heavy band on the origin were detected on the TLC plate from the reaction product of the acid procedure. Using Pasteur pipettes plugged with glass wool and a small layer of anhydrous sodium sulfate, the FAME were recovered from the silica gel using chloroform as solvent. Under a stream of N2, the chloroform was evaporated, and the FAME were transferred into 2 mL Agilent autosampler vials (Chromatographic Specialities Inc., Brockville, ON, Canada) using 1 mL iso-octane for gas chromatography (GC) analysis.

A gas chromatograph (Model 6890N; Agilent, Palo Alto, CA) used was equipped with a flame ionization detector operated in the splitless mode and flushed 0.3 min after injection, an autosampler (Model 7683B; Agilent), a 100-m CP-Sil 88 fused capillary column (100 m by 0.25 mm by 0.39 μm; Varian Inc., Mississauga, ON), and a software program (Agilent ChemStation, version A.10; Agilent). Injector and detector temperatures were kept at 240°C. Hydrogen was used as carrier gas at a flow rate of 1 mL/min and for the flame ionization detector at 40 mL/min. The injection volume was 1 μL, which contained about 1 to 2 μg of FAME. All FAME mixtures were routinely analyzed using 2 temperature programs in series followed by comparing these results to identify most FA isomers as previously described (Kramer et al., 2008). The FAME were identified by comparison with published separations (Kramer et al., 2002, 2008; Cruz-Hernandez et al., 2004) and GC reference FAME mixture number 463, spiked with the 4 positional CLA isomers mixture number UC-59M, and long-chain saturated FAME 21:0, 23:0, and 26:0 (Nu-Chek Prep, Inc., Elysian, MN).

Calculations and Statistical Analysis

Calculations for AID and ileal flow of dietary nutrients were as described by Nyachoti et al. (1997). The AID values and ileal flows were expressed as a percent and milligrams per kilogram DM intake, respectively. The sum of FA represents the sum of all FA within a specific class, such as SFA, MUFA, CLA, n-3 PUFA, and n-6 PUFA. Statistical analyses were performed using analysis of variance according to the MIXED model procedure (SAS Inst. Inc., Cary, NC) with the pig within the block as random effect and dietary treatment as fixed effect. The effect of litter was deemed not significant (P > 0.10) and was not included in the statistical model. Regression analyses using the GLM procedure of SAS were conducted to evaluate linear relationship between FA and AID. Differences among treatments means were assessed using the Tukey-Kramer honestly significant difference test of SAS. Probability levels less than P < 0.05 were considered significant, 0.05 < P < 0.10 was considered a trend, and P > 0.10 was considered not significant.


RESULTS

In general, pigs appeared healthy and readily consumed assigned feed allowances, and no abnormalities in animal behavior were observed. However, data from 5 pigs, 1 pig on R1 and 2 pigs each on R2 and R3, were removed from the study because insufficient amounts of digesta were obtained at slaughter.

Chemical Composition of Experimental Diets

The content of key nutrients in the diets is presented in Table 1. The analyzed contents of CP, Ca, P, and Na in the experimental diets were similar to anticipated values, indicating that diets were prepared properly.

The FA compositions reported in Table 2 represent the average of samples taken at the beginning and the end of the experiment. Differences in the contents of 9c-18:1, 18:2n-6, and 18:3n-3 and sum of all n-3 PUFA of feed samples taken at the beginning and end of the experiment varied by less than 3% of total FA content, indicating that auto-oxidation of unsaturated FA during storage of feed was minimal.


View Full Table | Close Full ViewTable 2.

Fatty acid (FA) content of the experimental diets (mg/kg DM)1

 
Item Grower flaxseed Grower control Finisher 1 control Finisher 2 control Finisher 2 flaxseed
Sum of all analyzed FA 50,271 33,616 39,897 34,916 40,577
SFA
    12:0 14.4 13.5 15.9 9.2 6.4
    14:0 54.0 287 442 239 30.3
    16:0 4,431 5,666 6,920 5,422 3,765
    18:0 1,869 2,392 3,346 2,040 1,287
    20:0 102 91.5 105 108 95.0
    22:0 89.4 55.6 52.0 54.0 62.9
    24:0 56.4 45.4 47.3 50.9 50.3
    Σ SFA 6,755 8,734 11,174 8,084 5,382
Branched-chain FA (BCFA)
    15:0 iso 15.0 12.6 21.8 11.2 4.40
    15:0 anteiso 2.27 17.8 29.0 15.3 1.34
    16:0 iso 4.92 13.9 22.8 11.9 4.72
    17:0 iso 1.59 8.1 12.7 6.8 0.88
    17:0 anteiso 12.9 44.1 66.3 39.3 10.0
    18:0 iso 2.32 12.3 18.5 10.9 1.35
    Σ BCFA 60.6 182 283 159 36.5
Trans-MUFA
    6t-8t-16:1 4.76 24.2 38.2 20.5 2.63
    9t-16:1 1.02 3.27 3.89 2.30 0.50
    10t-16:1 0.00 1.41 0.83 0.67 0.00
    11t/12t-16:1 0.00 2.56 3.75 2.16 0.00
    13t-16:1 12.9 44.1 66.3 39.3 10.0
    14t-16:1 6.01 6.24 5.49 4.41 2.22
    4t-18:1 1.06 1.63 1.90 1.41 1.44
    5t-18:1 1.32 1.75 2.39 1.63 0.89
    6t-8t-18:1 11.0 31.7 46.5 25.8 8.1
    9t-18:1 18.1 41.4 52.5 31.3 11.8
    10t-18:1 28.1 162 256 138 19.0
    11t-18:1 13.3 97.9 148.1 75.1 6.31
    12t-18:1 4.69 16.3 24.8 12.3 3.06
    Σ trans-MUFA 97.2 406 598 323 61.6
Cis-MUFA
    9c-12:1 12.1 0.72 1.44 0.86 2.73
    9c-14:1 1.9 37.0 61.2 32.7 0.68
    9c-16:1 33.2 160 232 130 21.3
    9c-17:1 23.0 61.0 93.4 56.0 17.7
    9c-18:1 10,612 9,133 11,378 9,571 9,047
    11c-18:1 539 342 403 289 430
    11c-20:1 94.9 105 124 103 80.6
    13c-22:1 11.4 7.50 8.38 7.60 6.72
    11c-24:1 8.46 6.53 6.54 5.35 5.26
    13c-24:1 4.35 11.4 10.7 9.27 8.91
    15c-24:1 4.66 7.06 6.99 5.54 4.30
    Σ cis-MUFA 11,598 10,191 12,765 10,478 9,785
n-6 PUFA
    16:2n-6 22.4 2.54 3.33 1.88 1.54
    18:2n-6 15,031 12,979 13,864 14,891 14,279
    18:3n-6 5.58 4.14 5.14 4.46 3.58
    20:2n-6 15.7 22.6 29.7 19.5 12.0
    20:3n-6 0.00 13.2 17.4 11.5 0.00
    20:4n-6 0.00 13.8 18.0 10.2 0.00
    22:2n-6 0.00 3.11 3.40 3.18 0.00
    22:4n-6 0.00 10.5 11.5 10.4 0.00
    22:5n-6 0.00 1.01 1.62 1.10 0.00
    Σ n-6 PUFA2 15,075 13,051 13,954 14,953 14,296
    Σ n-6 HUFA3 21.3 68.4 87.2 60.4 15.6
n-3 PUFA
    16:2n-3 3.42 2.39 3.19 2.74 2.47
    18:3n-3 16,481 878 745 720 10,835
    18:4n-3 0.00 0.38 0.57 0.96 0.00
    20:3n-3 12.99 4.13 4.73 3.36 8.24
    20:4n-3 0.00 1.92 3.08 1.89 0.00
    20:5n-3 0.00 5.42 7.47 4.99 0.00
    21:5n-3 0.00 2.84 2.39 2.59 0.00
    22:3n-3 13.3 24.8 32.3 21.2 12.0
    22:5n-3 0.00 5.40 8.18 4.73 0.00
    22:6n-3 0.00 3.80 5.48 3.29 0.00
    Σ n-3 PUFA4 16,511 929 812 766 10,857
    Σ n-3 HUFA5 26.3 48.7 64.3 43.0 20.2
    n-6 PUFA/n-3 PUFA 0.91 14.1 17.2 19.5 1.32
CLA
    9c11t-CLA 20.9 18.6 67.6 23.1 22.2
    8t10c-CLA 6.38 2.61 4.82 2.09 6.90
    9t11c-CLA 12.9 7.49 15.0 7.77 7.96
    10t12c-CLA 7.89 3.53 7.63 4.63 7.17
    11t13c/9c11c-CLA 28.5 2.20 4.37 5.08 27.5
    12t14c/13t15c-CLA 11.4 5.53 9.79 1.41 11.8
    11c13c-CLA 20.6 4.21 5.40 2.70 12.6
    12t14t/13t15c-CLA 28.6 44.8 136 74.2 25.3
    11t13t/13t15t-CLA 7.27 4.52 11.7 4.12 6.60
    9t11t/10t12t-CLA 29.6 11.6 25.7 9.13 31.1
    Σ CLA 174 105 288 134 159
1Diets were fed over these BW ranges: adaptation, up to 25 kg; grower, 25 to 50 kg; finisher 1, 50 to 85 kg; and finisher 2, 85 kg to 110 kg.
2n-6 PUFA is the sum of all n-6 PUFA.
3HUFA = highly unsaturated FA; n-6 HUFA is the sum of all n-6 PUFA minus 16:2n-6 and 18:2n-6.
4n-3 PUFA is the sum of all n-3 PUFA.
5n-3 HUFA is the sum of all n-3 PUFA minus 16:2n-3 and 18:3n-3.

The FA profile of ground FS was high in unsaturated FA such as 9c-18:1 (20.1%), 18:2n-6 (15.8%), and 18:3n-3 (51.5%), low in SFA (<10%), and contained trace amounts of trans FA isomers (<0.10%) whereas the FA composition of beef tallow included in the CON diets was high in SFA (40.4%) and 9c-18:1 (35.2%) and low in 18:2n-6 (9.29%) and 18:3n-3 (1.20%), with substantial amounts of trans FA isomers (4.06%). The inclusion of FS in the diets considerable increased the amount of 18:3n-3 in the diets, which represented 33.2 and 27.0% of total FA analyzed in the grower and finisher diets, respectively. There was a corresponding decrease in the amount of most SFA (14:0, 16:0, and 18:0), branched-chained fatty acids (BCFA; 15:0 to 18:0, iso or anteiso or both), trans- and cis-MUFA (except 9c-18:1), and n-6 and n-3 highly unsaturated FA (HUFA; all n-6 and n-3 PUFA, excluding 18:2n-6 and 18:3n-3, respectively) in FS diets as compared with CON diets (Table 2). On the other hand, 18:3n-3 content in the CON diets represented only 2.60 and 2.06% of total FA analyzed in the grower and finisher diets, respectively, with trace amounts of n-3 HUFA. The objective to create clear differences in the FA composition of the experimental diets was therefore achieved.

The sum of analyzed FA content for each diet was correlated to the analyzed crude fat content. The latter was consistently 1% unit greater than the calculated content for all diets. This systematic increase reflects an underestimation of the crude fat content of some of the main ingredients or a systematic bias in analytical procedures. This discrepancy did not influence the relative response to treatments.

Apparent Digestibility of Fat and Fatty Acids

The AID of crude fat and selected FA are presented in Table 3. The AID of crude fat and individual FA did not differ between R1 and R3, both of which represented feeding FS free diets before digesta sampling, and therefore, the data from these feeding regimens were pooled. This resulted in only 2 dietary treatments (FS and CON). No treatment effect was observed for AID of crude fat and the sum of selected FA listed in Table 3. The AID of SFA and trans- and cis-MUFA was similar for CON and FS. Myristic acid (14:0) was the only SFA to have a greater AID for CON than FS (P < 0.01) although all values were numerically greater in CON than FS. In general, the AID values of SFA decreased with increasing chain length, and a linear relationship was observed between chain length and AID values of SFA (P < 0.05) for both FS and CON. The AID of the sums of trans- and cis-MUFA were not affected by dietary treatment. However, the AID of the trans-18:1 isomers, 6t-8t- to 12t-18:1, were greater for CON than FS (P < 0.05). There were no treatment effects on AID of 18:2n-6 and 18:3n-3, but negative digestibilities were observed for most of the n-3 and n-6 HUFA metabolites and 9c11t-CLA. Only the AID value for 20:3n-3 was reported.


View Full Table | Close Full ViewTable 3.

Apparent ileal digestibilities (%) of crude fat and fatty acids (FA) in growing pigs (110 kg BW) fed diets containing 6% flaxseed or 1% tallow (Control)1

 
Item Flaxseed Control SEM P-value
n 7 8
Crude fat, % 75.6 73.9 3.4 0.568
Σ FA, % 79.4 81.8 2.8 0.623
SFA, %
    12:0 70.7 82.2 5.2 0.131
    13:0 86.1 90.6 2.8 0.253
    14:0 60.7 91.6 6.3 0.003
    16:0 70.7 76.6 5.4 0.429
    18:0 58.5 70.1 7.3 0.254
    20:0 70.9 78.5 5.8 0.352
    26:0 21.8 37.0 15.1 0.476
    Σ SFA, % 71.4 74.7 5.4 0.568
MUFA, %
    6t-8t-18:1 88.3 94.6 1.7 0.016
    9t-18:1 69.7 83.0 5.0 0.073
    10t-18:1 79.4 94.3 4.9 0.043
    11t-18:1 56.7 90.4 9.9 0.022
    12t-18:1 –24.6 76.4 27.2 0.018
    Σ trans-MUFA, % 76.4 87.9 7.8 0.198
    9c-12:1 62.3 29.2 11.0 0.042
    9c 14:1 85.1 97.2 5.0 0.092
    9c 16:1 77.9 88.3 2.8 0.069
    9c 18:1 83.3 88.2 3.2 0.262
    11c 18:1 75.9 64.5 7.6 0.296
    14c/16t-18:1 64.4 77.7 6.9 0.171
    15c-24:1 85.4 87.5 4.9 0.207
    Σ cis-MUFA, % 86.9 85.4 4.0 0.391
PUFA, %
    18:2n-6 80.2 86.1 0.9 0.284
    18:3n-3 89.8 86.7 2.4 0.370
    20:3n-3 –79.5 5.5 33.6 0.088
1Apparent ileal digestibility (AID) of crude fat and FA were calculated as AID = 100 – [100 × ([FA] digesta × [Marker] feed)/([FA] diet × [Marker] digesta)].

Ileal Flows of Fat and Fatty Acids

Ileal flows of crude fat and FA are presented in Table 4. There were no treatment effects on the content of crude fat in ileal digesta DM and on ileal flow of crude fat and the sum of all FA. Similarly, for SFA, BCFA, trans-MUFA, cis-MUFA, and n-6 PUFA, the ileal flows of individual FA and sums of FA were not affected by treatment, except for myristoleic (9c-14:1) and palmitoleic (9c-16:1), which were greater for CON than FS (P < 0.05). On the other hand, the ileal flow of individual and the sum of n-3 PUFA were greater for FS than CON (P < 0.05), with exception of docosatrienoic acid (22:3n-3), docosapentaenoic acid (22:5n-3), and docosahexaenoic acid (22:6n-3). Among the CLA isomers, the ileal flows of 9c11t-CLA and the sum of CLA were greater for FS than CON (P < 0.05) whereas ileal flow of 12t14c-CLA was lower for FS than CON (P < 0.01).


View Full Table | Close Full ViewTable 4.

Crude fat and fatty acid (FA) flow at the distal ileum in growing pigs (110 kg BW) fed diets containing 6% flaxseed or 1% tallow (Control)1

 
Item Flaxseed Control SEM P-value
n 7 8
Crude fat,2 % of DM 1.33 1.42 0.42 0.889
Fat flow g/kg DMI 15.4 12.6 2.0 0.689
Fatty acid flow, g/kg DMI 8.33 6.17 1.07 0.457
SFA, mg/kg DMI
    12:0 1.89 1.64 0.37 0.625
    14:0 14.5 20.0 3.7 0.294
    16:0 1,377 1,271 297 0.799
    18:0 626 610 129 0.929
    20:0 27.7 23.3 5.8 0.588
    22:0 36.5 29.7 7.3 0.506
    24:0 49.1 41.5 10.0 0.589
    Σ SFA 2,187 2,053 303 0.846
Branched-chain FA (BCFA), mg/kg DMI
    15:0 iso 0.71 0.55 0.27 0.672
    15:0 anteiso 0.47 0.72 0.11 0.108
    16:0 iso 3.39 5.97 1.50 0.232
    17:0 iso 0.41 0.84 0.16 0.078
    17:0 anteiso 6.09 7.63 1.74 0.530
    18:0 iso 0.88 1.68 0.33 0.098
    Σ BCFA 14.7 21.6 3.2 0.359
MUFA, mg/kg DMI
    5t-18:1 0.20 0.23 0.08 0.845
    6t-8t-18:1 0.95 1.39 0.19 0.111
    9t-18:1 3.57 5.31 0.95 0.206
    10t-18:1 3.92 7.88 1.35 0.052
    11t-18:1 4.50 7.22 1.23 0.131
    12t-18:1 3.81 2.91 1.10 0.561
    Σ trans-MUFA 27.1 39.9 8.1 0.569
    9c-12:1 1.03 2.36 1.14 0.409
    9c-14:1 0.26 0.90 0.15 0.008
    9c-16:1 18.9 115 21 0.043
    9c-18:1 1,508 1,127 281 0.340
    11c-18:1 104 103 24 0.976
    11c-20:1 23.5 21.5 5.1 0.776
    13c-22:1 3.70 2.94 0.71 0.449
    15c-24:1 10.4 10.6 2.3 0.952
    Σ cis-MUFA 1,803 1,554 226 0.568
n-6 PUFA, mg/kg DMI
    16:2n-6 0.37 2.19 1.52 0.399
    18:2n-6 2,829 2,067 560 0.339
    18:3n-6 2.23 2.44 0.58 0.794
    20:2n-6 15.3 19.1 4.6 0.551
    20:3n-6 10.3 16.9 4.3 0.288
    20:4n-6 95.0 170 39 0.176
    22:2n-6 2.83 3.72 0.90 0.481
    22:4n-6 23.5 56.2 11.7 0.063
    Σ n-6 PUFA 2,978 2,338 408 0.421
n-3 PUFA, mg/kg DMI
    16:2n-3 0.26 0.17 0.09 0.702
    18:3n-3 1,103 96 163 <0.001
    18:4n-3 0.64 0.02 0.08 <0.001
    20:3n-3 14.8 3.18 2.33 0.004
    20:4n-3 1.16 0.28 0.16 0.001
    20:5n-3 17.9 4.32 2.57 0.002
    22:3n-3 3.14 2.28 0.70 0.384
    21:5n-3 2.80 1.32 0.48 0.041
    22:5n-3 44.2 29.4 7.8 0.189
    22:6n-3 1.33 0.77 0.23 0.100
    Σ n-3 PUFA 1,190 137 177 <0.001
CLA isomers, mg/kg DMI
    9c11t-CLA 62.2 17.6 10.1 0.006
    8t10c-CLA 1.61 1.51 0.25 0.141
    9t11c-CLA 0.64 0.17 0.22 0.139
    11c13t-CLA 0.33 0.32 0.08 0.875
    10t12c-CLA 0.42 0.18 0.15 0.276
    12t14c-CLA 0.20 0.81 0.18 0.029
    9c11c-CLA 0.67 0.68 0.19 0.976
    11c13c-CLA 0.27 0.53 0.20 0.190
    12t14t-CLA 7.30 8.07 2.09 0.786
    11t13t-CLA 0.82 1.10 0.30 0.437
    9t11t/10t12t-CLA 2.04 2.41 0.70 0.699
    Σ CLA 76.5 48.4 9.5 0.095
1Total distal ileal flow (mg/kg DMI) was calculated as flows: FA flow = [FA] digesta × ([Marker] feed)/([Marker] digesta); Nyachoti et al. (1997).
2Analyzed crude fat content in digesta samples.


DISCUSSION

In several previous experiments, the nutritional value of FS meal was investigated in terms of the AID of CP, crude fat, and AA, but AID of FA was less frequently investigated (Eastwood et al., 2009). In the current study, attempts were made to measure FA contents in diets and digesta using improved analytical methods to ensure the complete and quantitative determination of most FA. Cruz-Hernandez et al. (2004, 2006) suggested that there is no single methylation procedure that adequately addresses the shortcomings of using either base- or acid-catalyzed methylation procedures, which certainly applies to the complex lipid mixtures in digesta. For this reason a base-catalyzed methylation was used to prevent the isomerizing of cis/trans- to trans/trans-CLA isomers and methoxy artifacts that generally occurs under acid-catalyzed conditions (Kramer et al., 1997). Because FFA, amides, and alk-1-enyl ether (plasmalogenic lipids) are not methylated under base conditions, acid-catalyzed methylation was also conducted to ensure the complete conversion of all lipids. In addition, complementary GC temperature programs were used to analyze most the FA and their isomers without prior silver-ion chromatographic separation (Kramer et al., 2008).

As expected, the grower and finisher diets containing ground FS differed substantially in the content of sum of all SFA, BCFA, n-3 PUFA, and trans-MUFA from CON diets, with little differences for cis-MUFA and n-6 PUFA (Table 2). This was largely the result of adding FS characterized by low SFA and high n-3 PUFA. Only trace amounts of total trans MUFA were present in ground FS because FS was not deodorized, a process known to cause formation of trans isomers (Ackman et al., 1974; Čmolík et al., 2000). To supply similar digestible Lys to DE intake ratios across diets, a small amount of beef tallow was added to all the CON diets, which resulted in increased levels of trans-FA (mainly 11t-18:1), BCFA, and CLA (mainly 9c11t-CLA), characteristic of fats from ruminants (Jensen, 2002). Trace amounts of CLA and trans FA were present in the FS diets, but their isomer distribution was random because it was produced by nonenzymatic processes (Juanéda et al., 2003; Azizian and Kramer, 2005).

The current study shows that the AID of the SFA decreased with increasing chain length, which is consistent with previous reports (Cera et al., 1988, 1989), whereas the digestibility of PUFA increased with the degree of FA unsaturation as previously noted by Jørgensen et al. (2000). The effect of increased unsaturation is consistent with the greater digestibility observed for 18:3n-3 than 18:2n-6 in the current study. It is of interest to note that the AID of the sum of all FA was greater than the AID observed for crude fat (Table 3), which was also observed by Jørgensen et al. (1992, 2000) and Duran-Montgé et al. (2007). These results indicate that other fat soluble components might have lower digestibility than FA.

The observed AID of FA in the CON diet, which included 1% beef tallow, were similar to the range in values reported by Jørgensen et al. (1992),involving a diet containing 1% animal fat and Overland et al. (1994) who used diets containing 6% rendered fat, 88.0 to 91.9% for 14:0, 69.7 to 80.4% for 16:0, 89.8 to 97.4% for 9c-16:1, 80.3 to 73.3% for 18:0, 79.0 to 90.4% for 9c-18:1, and 48.0 to 93.7% for 18:2n-6. Duran-Montgé et al. (2007) reported AID values for diets containing 10% beef tallow, and they were similar to those in the current study for 18:2n-6 (87.2%) and 18:3n-3 (88.7%). Htoo et al. (2008) reported decreased AID for 18:3n-3 (75.4%) in an experimental diet that included 30% of a 50:50 mixture of ground FS and field pea. In contrast, Duran-Montgé et al. (2007) reported greater AID for 18:3n-3 (98.6%) when 10% linseed oil was included in a barley-based diet. Several factors have been identified that result in variation in the AID of FA, including dietary fat level, fat source, losses of endogenous FA from the host into the gut (Jørgensen et al., 1993), levels of fiber, and other components in the diet and even analytical procedures (Bakker et al., 1995; Mroz et al., 1996; Wilfart et al., 2007).

The greater AID observed for CON than FS diets for myristic acid (14:0) and some MUFA (9c-12:1, 9c-14:1, and 9c-16:1) can be attributed to the content of mucilage, cyanogenic glycosides, goitrogens, and allergens present in FS (Bhatty, 1993), which can reduce FA digestibility. Changes in fat digestibility as a function of dietary fiber level have been well documented (Noblet and Perez, 1993; Bakker et al., 1995; Mroz et al., 1996; Wilfart et al., 2007). It is unclear, however, why pigs on FS had lower AID only for those FA and not for the main FA in the FS diet such as 9c-18:1, 18:2n-6, and 18:3n-3.

Regardless of fat source (such as fish oil, linseed oil, tallow, sunflower, rapeseed, and coconut oil), Jørgensen et al. (2000) and Duran-Montgé et al. (2007) observed low or negative AID values for stearic acid (18:0). Those researchers attributed the low AID of 18:0 to its formation from biohydrogenation processes of PUFA, specifically 18:2n-6 and 18:3n-3 in the intestinal lumen. Even though microbiota can alter to some extent the FA profile at the distal ileum, biohydrogenation rates of PUFA are generally greater in the hindgut (Jørgensen et al., 2000; Duran-Montgé et al., 2007). The differences in AID of 18:0 between the current results and those reported by Jørgensen et al. (2000) and Duran-Montgé et al. (2007) might be related to the methodology used to quantify the FA content of the digesta samples. Calcium in the small intestine will decrease FA absorption by increasing the formation of FA soaps (Atteh and Leeson, 1983). The solvent extraction method used by Duran-Montgé et al. (2007), without prior hydrolysis of lipids and acidification of the soaps to form FFA, may have underestimated the content of SFA from the digesta. The latter would lead to an overestimation of the AID of SFA.

The interpretation of ileal flows is of particular interest for the long-chain metabolites of the essential FA because they were virtually absent in the experimental diet. Even though n-3 and n-6 HUFA were not detected in FS diets, small amounts of n-3 and n-6 HUFA, such as 20:4n-6, eicosapentaenoic acid (20:5n-3), 22:5n-3, and docosahexaenoic acid (22:6n-3), were observed in the ileal flow, which indicates activity of the Δ6- and Δ5-desaturases and chain-elongases in the upper gut of pigs. It remains to be resolved whether the microbiota in the intestinal lumen can elongate and desaturate 18:2n-6 and 18:3n-3 and to what extent microbial fermentation in the upper gut contributes the overall HUFA content in the digesta of nonruminant animals. The contribution of endogenous FA losses from the host to the ileal flow of these FA should be considered as well.

The ileal flow of CLA was substantially greater when FS was included in the diet compared with CON (Table 4), even though only the CON diet contained beef tallow, a source of CLA. In addition, 9c11t-CLA was the predominant CLA isomers in the ileal flow of the FS group, even though the trace amount of CLA present in the ground FS consisted of a random distribution of CLA isomers (Juanéda et al., 2003; Azizian and Kramer, 2005). This increase of 9c11t-CLA could only be attributed to its biosynthesis from 18:2n-6 by the action of Butyrivibrio fibrisolvens or other bacteria capable of this biohydrogenation process. Alternatively, 9c11t-CLA could be de novo synthesized from 11t-18:1 by Δ9-desaturase (Griinari et al., 2000). Both the synthesis of 9c11t-CLA from 18:2n-6 by bacteria such as Butyrivibrio fibrisolvens and the conversion of 11t-18:1 to 9c11t-CLA by Δ9 desaturases in ruminants has been well established (Bauman and Griinari, 2003). Chin et al. (1994) and Adlof et al. (2000) suggested that rats and humans, respectively, can also synthesize CLA isomers in the upper gut. Regardless of how 9c11t-CLA was synthesized in the gut of pigs, it was found to be greater in pigs fed FS than when fed CON (Table 4). This indicates that feeding ground FS stimulated CLA synthesis from 18:2n-6 or its Δ9 desaturation from the 11t-18:1 precursor. The dietary level of 18:2n-6 was not greater in FS than CON diets but possibly more available. It is of interest to note that the CLA isomer profile of CON was similar in the diet and ileal flow, which was not surprising considering that both are produced in biological systems. These results indicate that further investigationsare necessary to determine the extent of CLA synthesis in the small intestine of pigs and evaluate its contribution to the host animal.

In summary, the current study indicated that type of fat from either ground FS or tallow included in corn–wheat–soybean based diets does not affect the AID of the sum of FA and total fat in growing pigs. The supplemental dietary FA from ground FS (6%) was highly digestible (<80%) and greater than reported values using 15% of ground FS. The addition of 1% tallow resulted in similar AID values reported in the literature. The FA digestibility decreased with chain length and increased with increasing degree of unsaturation of FA. It appears that pigs are able to synthesize CLA isomers, in particular 9c11t-CLA, and HUFA in their upper gut from their dietary 18:2n-6 and 18:3n-3 precursors. Further studies are needed to quantify the extent of CLA isomers and HUFA synthesis in the small intestine of nonruminant species, specifically the 20:5n-3, 22:5n-3, and 22:6n-3, and to what extent these HUFA are absorbed. The contribution of endogenous FA losses to the ileal flow should also be considered.

 

References

Footnotes


Comments
Be the first to comment.



Please log in to post a comment.
*Society members, certified professionals, and authors are permitted to comment.