Effects of Soybean Oil or Whole Cotton Seed Addition on Accumulation of Conjugated Linoleic Acid in Beef of Fattening Brahman × Thai-Native Cattle

Effects of soybean oil or whole cotton seed addition on conjugated linoleic acid (CLA) and performance of fattening Brahman×Thai-Native cattle were studied. Eighteen fattening cattle averaging 241±24 kg body weight and approximately 1 year old were stratified by live weight into three groups and randomly assigned by group to one of three dietary treatments. The treatments were control (concentrated 14% crude protein), control and supplemented with 170 g/d soybean oil, control plus 170 g/d of oil from whole cotton seed. All animals were weighed before and after the experimental period and 4 cattle per treatment were randomly slaughtered then carcass measurements were obtained. There were no statistically significantly differences in the final body weight, average daily gain and dry matter intake among treatments. However, the crude protein intake was significantly decreased (p<0.01) when whole cotton seed was fed compared with control and soybean oil treatments. The carcass composition and carcass characteristics were not significantly different in Longissimus and Semimembranosus muscle by feeding soybean oil and whole cotton seed compared with the control treatment. Supplementation of soybean oil increased (p<0.01) cis-9, trans-11 CLA by 116% in Longissimus muscle and by 240% in Semimembranosus muscle. However, whole cotton seed did not increase cis-9, trans-11 CLA in both muscles. The present study successfully increased cis-9, trans-11 CLA content of muscle lipids by soybean oil but not by whole cotton seed. (

Researchers have successfully increased the cis-9, trans-11 CLA content of muscle lipids by plant oils (Eagle et al., 2000;Mir et al., 2002Mir et al., , 2003;;Choi et al., 2006;Wang et al., 2006;Noci et al., 2007) and oil seeds (Bolte et al., 2002).Thus, supplementation of fat sources rich in linoleic acids such as plant oils or oil seeds may increase CLA content in muscle lipid.However, no comparison between soybean oil (SBO) and whole cotton seed (WCS) has been made in previous researches published.
Brahman×Thai-Native cattle are widely raised by Thai farmers and accounted for 40% of total beef cattle population of 8.8 million heads (Department of Livestock Development, 2007).Twenty percent of these cattle are raised for yearling cattle which are used as grilled beef in restaurants.The requirement of good quality beef for grilled beef is that cattle will be fattened for a short period of 3 to 4 mo and body weight at the start of fattening period is approximately 240-250 kg and the final weight of finishing stage is between 300 to 330 kg.
The objective of the present study was to determine the effect of whole cotton seed (WCS) or soybean oil (SBO) supplementation on CLA accumulation in beef and on carcass characteristics of young growing fattening cattle.

Animals and feeding
Experiment was conducted in accordance with the principles and guidelines approved by the Suranaree University of Technology Animal Care and Use Committee which followed Guidelines for the Care and Use of Agricultural Animals in Agricultural Research and Teaching (1 st revised edition, 1999; Association Headquarters, 1111 North Dunlap Avenue, Savoy, IL 61874).Eighteen crossbred Brahman beef bulls, averaging 241±24 kg of body weight and approximately 1 year of age, were stratified by body weight into three groups and randomly assigned by group to one of three dietary treatments.The treatments were control (commercial concentrate 14% crude protein; CP), control plus 170 g/d of SBO and control plus 170 g/d of oil from WCS. Ingredient and nutrient compositions of concentrate used in the experiment are given in Table 1 and Table 2 respectively.Cattle were individually housed and fed 3.5 kg/d of concentrate divided in two equal meals at 0800 and 1600 h and ad libitum rice straw and free access to clean water.Soybean oil and whole cotton seed were top-dressed on concentrates at each feeding.The experiment lasted for 109 d.Fasted live weights were recorded at the start of the trial and then every two weeks throughout the experimental period.At the end of the experiment, cattle were weighed and 4 cattle per treatment were randomly sampled and then slaughtered.

Sample collection and analysis
Feed offered and left after consuming of individual cattle were weighed and collected on two consecutive days of each period of 14 d.Samples were taken and dried at 60°C for 48 h.Feed intakes were then determined by the difference between feed offered and left after eating.At the end of the experiment, feed samples were performed on well-mixed subsamples and then taken for further chemical analysis.Samples were ground through 1 mm sieve and analyzed for proximate (AOAC, 1990) and detergent analyses (Van Soest et al., 1991).

Carcass collection and analysis
Muscle samples from 6 cattle per treatment were cut from outside Longissimus dorsi (LD) and Semimembranosus (SM) muscle on the left side of each carcass.All samples were placed in plastic bags and chilled on ice.At the laboratory, samples were chilled at 4°C for 48 h.Then measurements were made on Longissimus muscle area at 12th rib, color and shear force.Muscle samples were removed from the plastic bags and cut.These steaks were measured for meat color by Chroma meter (Minolta CO., LTD) and then L* (lightness), a* (redness) and b* (yellowness) value reading were made in six locations from each piece of meat.The LD muscle areas were measured (Delta-t Devices LTD.England).Shear force was done with    , 1996); NE M = Net energy for maintenance = 1.37ME-0.138ME 2 +0.0105ME 3 -1.12(NRC, 1996); NE G = net energy for gain = 1.42ME-0.174ME 2 +0.0122ME 3 -1.65 (NRC, 1996).

Fatty acid analysis
Feed and meat fat were extracted using a modified method used by Folch et al. (1957) and Metcalfe et al. (1966).Before the extraction, meat samples were thawed and each sample was chopped coarsely and blended in a blender machine.A 15 g of each sample was homogenized for 2 min with 90 ml of chloroform-methanol (2:1) (Nissel AM-8 Homoginizer, Nihonseikikaisha, LTD., Japan).Each sample was further homogenized for 2 min with 30 ml of chloroform.Then, each sample was separated with separating funnel and 30 ml of deionized water and 5 ml of 0.58% NaCl were added.The lower layer was removed and placed in a screw-cap test tube and stored at -20°C until methylation.
Fatty acid methyl esters (FAME) were prepared by the procedure described by Ostrowska et al. (2000).In this procedure, approximately 30 mg of the extracted oil was placed into a 15-ml reaction tube fitted with a teflon-lined screw cap.One and a half ml of 0.5 M sodium hydroxide in methanol was added.The tubes were flushed with nitrogen, capped, heated at 100°C for 5 min with occasional shaking and then cooled to room temperature.One ml of C17:0 internal standard (2.00 mg/ml in hexane) and 2 ml of boron trifluoride in methanol were added and heated at 100°C for 5 min with occasional shaking.After methylation was completed, 10 ml of deionized water was added.The solution was transferred to a 40-ml centrifuged tube and 5 ml of hexane was added for FAME extraction.The solution was centrifuged at 2,000 g, at 10°C for 20 min and then the hexane layer was dried over sodium sulfate and was taken into a vial to be analyzed by gas chromatography (GC) (Hewlett Packard GC system HP6890 A; Hewlett Packard, Avondale, PA) equipped with a 100 m×0.25 mm×0.2 μm film fused silica capillary column (SP2560, Supelco Inc, Bellefonte, PA, USA).Injector and detector temperatures were 240°C.The column temperature was kept at 70°C for 4 min, then increased at 13°C/min to 175°C and held at 175°C for 27 min, then increased at 4°C/min to 215°C and held at 215°C for 31 min.

Statistical analysis
All data were statistically analyzed as a completely randomized block design using ANOVA procedure of SAS (SAS, 1998).Differences between treatment means were statistically compared using Duncan's New Multiple Range Test (Steel and Torrie, 1980).

RESULT AND DISCUSSION
The chemical composition of experimental diets, whole cotton seed and rice straw are presented in Table 2. Soybean oil diet had higher fat content than other diets.Concentration of C18:2 was increased by supplementation of SBO or WCS in the diets.C18:3 was negligibly detected in SBO and WCS diets in the present study.
There were no significant differences in protein, lipid and moisture content of both Longissimus muscle (LM) and Semimembranosus muscle due to SBO or WCS supplementation (Table 5).Carcass characteristics including dressing percentage, Longissimus dorsi area, shear force, color and marbling score were also not statistically significantly different among treatment diets (Table 5).
Researchers have successfully increased cis-9, trans-11 CLA content of muscle lipids by various sources of oils (Engle et al., 2000;Mir et al., 2002Mir et al., , 2003;;Noci et al., 2007).CLA contents of muscle were increased by 45% when feeding 4% SBO (Engle et al., 2000), by 339% when feeding 6% SFO (Mir et al., 2002), by 30 or 75% when feeding 3 or 6% SFO (Mir et al., 2003) and by 144 or 73% when feeding SFO or linseed oil (LSO) respectively (Noci et al., 2007).However, some reports found no significant differences in CLA content in adipose or muscle tissues (Dhiman et al., 1999;Beaulieu et al., 2000;Dhiman et al., 2005).Increases in cis-9, trans-11 CLA accumulation in muscle by SBO can be attributed to the fact that SBO is rich in C18:2 which is used to promote direct synthesis of CLA.The biohydrogenation is incomplete in the rumen; CLA isomer and C18:1 trans-11 vaccenic acid are intermediates escaped from the rumen and then converted to produce CLA (C18:2 cis-9, trans-11) in tissue by the action of Δ 9 desaturase (Griinari et al., 1998;Baumam et al., 1999;Corl et al., 2001).In the present study, supplementation of WCS did not affect the CLA content in muscle although it is rich in C18:2 content.Palmquist et al. (1995) suggested that digestion and utilization of fatty acids in whole cotton seed and whole sunflower seed are dependent upon rumination and mastication to provide microbial access to seed contents.Page et al. (1997) expected that WCS depresses stearoylcoenzyme A desaturase activity in subcutaneous adipose tissue and liver due to its cyclopropene fatty acid content in WCS if fed for a sufficiently long period of time.Madron et al. (2002) suggested that possibility of the biohydrogenation is more complete in the rumen, resulting in the formation of more stearic acid and thus less C18:1 trans-11 vaccenic acid and CLA would escape the rumen.
There were significant decreases (p<0.001) in C12:0, C14:0, C14:1 and C15:0 in LM and SM when WCS diet was fed compared with control diet (Tables 6 and 7).Feeding SBO or WCS diet reduced (p<0.05)C16:0 and C16:1 in LM and C16:1 in SM.The present study confirms the result of Engle et al. (2000) who found a decrease in C16:1 but not in C16:0 in muscle and adipose tissues when steers were fed diet containing 4% SBO.Similar results were also reported with 5% SBO (Beaulieu et al., 2002), with 6% SFO (Mir et al., 2002), and with 3 or 6% SFO (Mir et al., 2003).However, Noci et al. (2007) indicated that C12:0, C14:0 and C16:0 were increased in muscle tissue when cattle were fed SFO and LSO.Dhiman et al. (2005) reported that C12:0-C16:0 in adipose and muscle tissues from steers fed 2 or 4% SBO were unaffected while C14:0 was increased and C16:1 was decreased.The reduction in C16:0 and C16:1 caused by SBO or other plant oils addition to the diets is probably due to the negative feedback inhibition of fatty acid synthesis by the exogenous fatty acids.
Stearic acid (C18:0) in LM and SM were similar in all diets.However, both LM and SM lipids showed a reduction in C18:1 when WCS was fed.Moreover, the addition of WCS decreased C18:2 in SM and C18:3 in LM.The soybean oil supplemented cattle showed similar C18:1, C18:2 and C18:3 in LM and SM to the control cattle.Other reports showed 10 and 12% increases in C18:0 when 5% SBO was supplemented (Beaulieu et al., 2002) and 10% increase in C18:0 when 4% SBO was fed (Dhiman et al., 2005).Griswold et al. (2003) reported a linear increase in C18:2, while C18:3 was unaffected when 0, 4 or 8% SBO were fed to steers.Mir et al. (2003) also found an increase in C18:2 when 3 or 6% SFO was added to diets.Recently, Noci et al. (2007) indicated that C18:1 and C18:2 were significantly increased while C18:0 was reduced by SFO and LSO supplementation.Short-and medium-chain fatty   supplementation, but not by WCS, however, both SBO and WCS had no effect on DMI, final body weight, and ADG.

Table 1 .
Ingredient composition of concentrate used in the trial

Table 3 .
Fatty acid compositions of experimental diets, soybean oil, whole cotton seed and rice straw

Table 4 .
Effects of SBO or WCS supplementation on feed intake and growth performance +SBO = Supplementation with 170 g/d soybean oil; +WCS = Supplementation with 170 g/d oil from whole cotton seed; SEM = Standard error of mean; BW = Body weight; ADG = Average daily gain.

Table 5 .
Effects of SBO or WCS supplementation on carcass composition and carcass characteristics +SBO = Supplementation with 170 g/d soybean oil; +WCS = Supplementation with 170 g/d oil from whole cotton seed; SEM = Standard error of mean.CONCLUSIONIt can be clearly concluded, in the present study, that the CLA content in beef was increased by SBO

Table 6 .
Effects of SBO or WCS supplementation on fatty acid composition of Longissimus muscle (mg/g fat) = Supplementation with 170 g/d soybean oil; +WCS = Supplementation with 170 g/d oil from whole cotton seed; SEM = Whole standard error of mean.

Table 7 .
Effects of SBO or WCS supplementation on fatty acid composition of Semimembranosus muscle (mg/g fat) Supplementation with 170 g/d soybean oil; +WCS = Supplementation with 170 g/d oil from whole cotton seed; SEM = Standard error of mean. 1 Fatty acids: <C 16:0 originated from de novo synthesis fatty acids; >C 16:0 were performed fatty acids.