Synthesis and Biohydrogenation of Fatty Acids by Ruminal Microorganisms In Vitr0 1 Z. WU and D. L. PALMQUIST2 Department of Dairy Science Ohio Agricultural Research and Development Center The Ohio State University Wooster 44691 ABSTRACT
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Abbreviation key: AV animal-vegetable blend fat, BH = biohydrogenation, CS = calcium soap, dpm = disintegrations per minute, FA = fatty acids, LCFA = long-chain fatty acids.
Ruminal degradation, synthesis, and biohydrogenation of fatty acids were examined in vitro. Diets were incubated with ruminal contents, and changes of fatty acids were measured Two fat supplements, a calcium soap and an animalvegetable blend, were included in diets at various levels. Addition to diets of acetate and isoacids (collective term for certain short-chain acids) also were tested for effects on fatty acid synthesis. Overall, 6.6 mg of fatty acids/g of fat-free diet were synthesized during 24-h incubation regardless of supplementations. Fatty acids synthesized in greatest amounts were odd-numbered or branched chains, whereas chains of 16 and 18 carbons changed little, and chains shorter than 14 carbons decreased. Degradation of [1_ 14C]palrnitic acid was negligible, as determined by recovery of the label in C02 (.03%) and acetate (1.09%) after 4-h incubation with rumen contents. Biohydrogenation of fatty acids averaged 47% in diets containing calcium soap and 71 % with animal-vegetable blend Synthesis and biohydrogenation were similar to those measured previously in vivo, showing that in vitro measurements reliably predicted metabolism of fatty acids in vivo. (Key words: fatty acid synthesis, biehydrogenation, ruminal microorganisms)
INTRODUCTION
Received January 22, 1991. Accepted April 5, 1991. ISalaries and research suppon provided by state and federal funds to the Ohio Agricultural Research and Development Center, 1be Ohio State University, and by Church and Dwight Co., Inc., Princeton, NJ. Manuscript Number 332-90. 2RepriDts .
1991 J Dairy Sci 74:3035--3046
Rumen microorganisms are known to synthesize long-chain fatty acids [LCFA; (10)], and LCFA are not degradable in the rumen because the anaerobic condition is not favorable for oxidation of fatty acids (FA). Thus, net synthesis should occur, causing the amount of FA leaving the rumen to be larger than that ingested. However, evidence of degradation has been reported from studies using isolated cultures of rumen bacteria or protozoa (6). Apparent decreases (30), as well as increases (25), of FA in the rumen have been reported from studies measuring FA in duodenal digesta of animals fitted with T-cannulas. It is not certain whether decreases resulted from ruminal degradation, absorption, or underestimation of duodenal flow. Acetic acid is the basic unit for FA synthesis, whereas certain other short-chain acids (isobutyric, isovaleric, 2-methylbutyric, and valeric acids, known as isoacids) can serve to initiate chain synthesis (1). Synthesis of FA by rumen bacteria may be influenced by FA present in media. Demeyer et al. (5) concluded that ruminal microbes are more likely to incorporate preformed FA into cell lipids than to synthesize new FA de novo. This was based on larger recovery of radioactivity in microbial lipids from radioactive linoleic acid than from labeled acetate or glucose when these were incubated with strained rumen fluid In the present study, a series of experiments was conducted in vitro to determine extent of
3035
3036
WU AND PALMQUIST
Collection of C02. Central plastic wells were suspended inside the incubation flasks from rubber serum stoppers closing the flasks. A folded sheet of filter paper soaked with 1 ml of ION NaOH solution was placed in the central well to absorb
=
Abbreviation key: AV animal-vegetable blend fat, BH = biohydrogenation, CS = calcium soap, dpm = disintegrations per minute, FA = fatty acids, LCFA = long-chain fatty acids.
Ruminal degradation, synthesis, and biohydrogenation of fatty acids were examined in vitro. Diets were incubated with ruminal contents, and changes of fatty acids were measured Two fat supplements, a calcium soap and an animalvegetable blend, were included in diets at various levels. Addition to diets of acetate and isoacids (collective term for certain short-chain acids) also were tested for effects on fatty acid synthesis. Overall, 6.6 mg of fatty acids/g of fat-free diet were synthesized during 24-h incubation regardless of supplementations. Fatty acids synthesized in greatest amounts were odd-numbered or branched chains, whereas chains of 16 and 18 carbons changed little, and chains shorter than 14 carbons decreased. Degradation of [1_ 14C]palrnitic acid was negligible, as determined by recovery of the label in C02 (.03%) and acetate (1.09%) after 4-h incubation with rumen contents. Biohydrogenation of fatty acids averaged 47% in diets containing calcium soap and 71 % with animal-vegetable blend Synthesis and biohydrogenation were similar to those measured previously in vivo, showing that in vitro measurements reliably predicted metabolism of fatty acids in vivo. (Key words: fatty acid synthesis, biehydrogenation, ruminal microorganisms)
INTRODUCTION
Received January 22, 1991. Accepted April 5, 1991. ISalaries and research suppon provided by state and federal funds to the Ohio Agricultural Research and Development Center, 1be Ohio State University, and by Church and Dwight Co., Inc., Princeton, NJ. Manuscript Number 332-90. 2RepriDts .
1991 J Dairy Sci 74:3035--3046
Rumen microorganisms are known to synthesize long-chain fatty acids [LCFA; (10)], and LCFA are not degradable in the rumen because the anaerobic condition is not favorable for oxidation of fatty acids (FA). Thus, net synthesis should occur, causing the amount of FA leaving the rumen to be larger than that ingested. However, evidence of degradation has been reported from studies using isolated cultures of rumen bacteria or protozoa (6). Apparent decreases (30), as well as increases (25), of FA in the rumen have been reported from studies measuring FA in duodenal digesta of animals fitted with T-cannulas. It is not certain whether decreases resulted from ruminal degradation, absorption, or underestimation of duodenal flow. Acetic acid is the basic unit for FA synthesis, whereas certain other short-chain acids (isobutyric, isovaleric, 2-methylbutyric, and valeric acids, known as isoacids) can serve to initiate chain synthesis (1). Synthesis of FA by rumen bacteria may be influenced by FA present in media. Demeyer et al. (5) concluded that ruminal microbes are more likely to incorporate preformed FA into cell lipids than to synthesize new FA de novo. This was based on larger recovery of radioactivity in microbial lipids from radioactive linoleic acid than from labeled acetate or glucose when these were incubated with strained rumen fluid In the present study, a series of experiments was conducted in vitro to determine extent of
3035
3036
WU AND PALMQUIST
Collection of C02. Central plastic wells were suspended inside the incubation flasks from rubber serum stoppers closing the flasks. A folded sheet of filter paper soaked with 1 ml of ION NaOH solution was placed in the central well to absorb
