INCLUSION OF PROTECTED FAT IN DIETS ON THE MILK PRODUCTION AND COMPOSITION OF SAANEN GOATS

According on the fat source included in the diets for goats, the milk production can be increased and milk fatty acid composition can be improved. This study was conducted to evaluate the inclusion of protected fat (CSFA – calcium salts of fatty acids) in the diets of lactating Saanen goats on milk production and composition. Five Saanen goats (63.23 ± 9.08 kg body weight and 105 ± 3 days of lactation) were distributed in a Latin square design (5 × 5) with five diets: control (without the addition of CSFA) and the other diets with: 6.25, 12.50, 18.75 and 25.0 g CSFA per kg of dry matter (g/kg DM), respectively. Dry matter intake (DMI) and milk production (MP) were evaluated. Milk samples were collected to determine milk composition and fatty acid profile. Feed efficiency (FE) was estimated. The supplementation CSFA up to 25 g/kg DM did not influence the DMI, MP and FE. Milk components production (g/d) was not influenced by the inclusion of CSFA in the diet; however, there was a linear reduction of 0.87 g protein per kg of milk for each 1% CSFA included in the diet. The concentration of linolenic acid (18:3n-3) in milk was improved with a maximum point of 24 g CSFA per kg DM. CSFA can be included in the diets of lactating Saanen goats up to 25 g/ kg DM with no interference on DMI and MP.


INTRODUCTION
Goat milk in Brazil is mainly consumed as an alternative feed for people that present with an allergy to milk from cows because it has a low αS1-casein content (Greppi;Roncada;Fortin, 2008;Tomotake et al., 2006).Furthermore, the milk from goats contains fatty acids that are essential for human health, and small globules of fat when compared to the milk from cows (Attaie;Richter, 2000) what check a better digestibility for goat milk.The use of goat milk as a functional feed indicates a need for the specialization of the production chain as well as an improvement in the management of sanitation, reproduction, milking, and feed.
Generally, lipid supplements are added to the diets of lactating females to increase energy density; however, this also contributes to other purposes such as improving fat-soluble vitamin absorption, supplying important fatty acids for tissue membranes, acting as a precursor of metabolism regulation, increasing the efficiency of fat deposition, and improving milk synthesis (Palmquist;Mattos, 2011).
Protected fat is one of the supplements that is available in the market and is an ion complex of calcium with long chain fatty acids whose raw material can be soybean oil or palm oil according to the commercial product.Protected fat is also called inert because it remains intact from the action of the rumen microorganisms without interfering in the fermentation process (Sirohi;Walli;Mohanta, 2010).Once the calcium ions are dissociated in the acidic conditions of the abomasum, the lipids become available for digestion and absorption directly from the intestine (Silva et al., 2007).
The dry matter intake can be reduced when a large percentage of protected fat is provided for goats (Sanz Sampelayo et al., 2002a); however, the efficient lipid digestion can increase milk production (Titi, 2011).Protected fat also allows for the increase of polyunsaturated fatty acid concentrations (Sanz Sampelayo et al., 2002b), which is a strategy to improve milk composition.
This study was conducted to evaluate the inclusion of calcium salts of fatty acids in diets on the dry matter intake, feed efficiency, milk production and composition, and concentration of fatty acids of milk from Saanen goats.

MATERIAL AND METHODS
The experiment was conducted at the Iguatemi Experimental Farm of the Universidade Estadual de Maringá, southern Brazil.Five multiparous Saanen goats (63.23 ± 9.08 kg body weight and 105 ± 3 days lactation) selected according to milk production in relation to body weight were distributed in a Latin square design (5 × 5) with five diets: control (without the addition of CSFAcalcium salts of fatty acids) and the other diets with: 6.25, 12.50, 18.75 and 25.0 g CSFA per kg dry matter (g/kg DM), respectively.
The ingredients that were utilized to prepare the diets were ground corn, soybean meal, CSFA (calcium salts of fatty acids -Lactoplus ® containing 1,760 g/kg total digestible nutrients, 820 g/kg ether extract, 100 g/ kg calcium, 260 g/kg oleic acid, and 420 g/kg linoleic acid), and mineral supplement ® .The oat hay (920 g/kg dry matter, 78 g/kg crude protein and 698 g/kg neutral detergent fiber) was uzed at a proportion of 570 g/kg DM (Table 1).The diets were adjusted to provide 700 g total digestible nutrients and 155 g crude protein per kg dry matter, respectively, according to the NRC (2007).The feed samples were processed in a Willey type mill using a 1 mm screen and stored in plastic bottle for further analyses.
Dry matter (DM), crude protein (CP), ash, and ether extract (EE) were analyzed according to AOAC as described by Silva and Queiroz (2002), and neutral detergent fiber was analyzed according to Van Soest, Robertson and Lewis (1991) as adapted by Souza et al. (1999).Organic matter was estimated by calculating the difference between dry matter and ash contents.
The total carbohydrates (TC) and total digestible nutrients (TDN) were estimated according to the equation described by Sniffen et al. (1992) Each experimental period was composed of 21 days, with 15 days for the animals to adapt to the diets and six days of data collection (week of evaluation), thereby totaling 105 days of experiment.The goats were housed in individual pens with ad libitum water, and access to the solarium in the morning.
Body weight was recorded at the beginning of each experimental period after the morning milking and before the supply of the ration.
Goats were fed at 9h and 16h and the orts were measured daily to determine the intake.Samples of orts were collected daily from 16th to 21th of the experimental periods.
The samples of orts were stored in a freezer, and at the end of the collections they were defrosted, dried in a forced oven for 72 hours at 55 o C, ground in a Willey type mill using a 1 mm screen, and analyzed for dry matter, according to the procedures previously described for feed samples, to determine the dry matter intake (DMI).
The goats were milked twice daily (7h30 and 15h30) followed by individual milk weighting.For each experimental period, individual milk samples were collected at the 16th day proportionally to milk production of the morning and afternoon milkings.
Another two milk samples were collected in plastic bottles without addition the preservative and frozen, one for milk fatty acid and the other one for urea analyzes.
Milk fat was obtained by centrifugation according to the methodology described by Murphy, Connolly and Mcneill (1995), and was esterified according to the method 5509 of ISO (1978) in a solution of KOH/methanol and n-heptane.Fatty acid methyl esters were quantified by gas chromatography (Trace GC Ultra, Thermo Scientific, USA) equipped with a flame ionization detector at 240°C and a capillary column of fused silica (100 m length, 0.25 mm internal diameter, and 0.20 μm, Restek 2560).
Fatty acids were quantified (g/100g lipids) by comparison with the retention time of the methyl esters from fatty acids of the sample standards tricosanoic acid methyl ester (23:0) (Sigma Aldrich ® , Brazil).The column parameters were as follows: initial column temperature of 65°C was maintained for 8 min; the temperature was then programmed at 50°C/min to 170°C; this temperature was maintained for 40 min and then increased 50°C/min to 240°C and remained for 28.5 min.Injector and detector temperatures were 220 and 245°C, respectively.The gas flow was 1.5 ml/min for hydrogen (carrier gas), 30 ml/ min for N2 (auxiliary gas), 35 ml/min for H2 and 350 ml/ min for compressed air.With a microliter syringe, 2 µL of samples were inject with a split ratio of 1:100.Fatty acid peaks were identified by comparison of the retention times of pure methyl ester standards (Sigma Aldrich ® , Brazil).
At the 21th day of the experimental periods blood samples were collected from the goats after the morning milking (before feeding) by jugular vein puncture using 5 mL vials.The samples were centrifuged (3,000 rpm for 15 minutes) to obtain the serum, which was stored in eppendorfs and analyzed by a biochemical semiautomatic analyzer using an enzyme-colorimetric kit (Gold Analisa Diagnostica Ltda ® ) for urea concentration.
Feed efficiency was calculated by determining the ratio between the average milk production and the average dry matter intake, in kg/kg.Ciênc. Agrotec., Lavras, v.39, n.2, p.164-172, mar./abr., 2015 The data were submitted to variance analysis via polynomial regression (P<0.05) by using the SAEG (Genetic and Statistical Analyses System -UFV version 7.1) program.

RESULTS AND DISCUSSION
Dry matter intake (DMI), milk production (MP), and milk production corrected for 35 g of fat per kg of milk (FCM 3.5% ) were not influenced (P>0.05) by the inclusion of CSFA in the diets, and had average values of 2.04, 2.07 and 1.9 kg/d, respectively (Table 2).
DMI determines the nutrient supply needed to meet the requirements for maintenance and production (Carvalho et al., 2006); therefore, among the several factors that determine milk production, maximization of the intake is one of the decisive factors that should be taken into consideration.The result observed for DMI indicates that although the protected fat presents with the characteristic odor of soap (fatty acid calcium soap), its inclusion in the diet did not restrict DMI, thereby leading to a good level of acceptance of the product without influencing the flavor for goats.
The use of CSFA for lactating goats in diets in proportions from 30 to 90 g/kg DM does not seems to influence the intake (Silva et al., 2007;Teh et al., 1994).However, when CSFA fat is included in greater proportions in the diet, the intake can be influenced.In a study with inclusion levels of 0, 90 and 120 g protected fat per kg of dry matter in the diets of goats, Sanz Sampelayo et al. (2002a) reported that there was a refusal in the intake of concentrate due to the reduced palatability of diets with protected fat.
There was no influence (P>0.05) of the inclusion of CSFA in diets on milk production.Sanz Sampelayo et al. (2002b) supplemented lactating goats with higher amounts of protected fat (0, 90 and 120 g/kg DM) and did not observe changes in the milk production.The authors justified this due to the similar metabolizable energy intake results among the evaluated diets, since the energy of the diet is an important factor that limits milk production.
Feed efficiency (FE), and feed efficiency considering the milk production corrected (FEc) with averages of 1.01 and 0.95, respectively, were not influenced due the results obtained for milk production and dry matter intake.In a study conducted where lactating Saanen goats received different roughage:concentrate proportions, Zambom et al. ( 2005) observed a value of 1.05 for the FE, when the goats were provided with 60% oat hay, a value that is close to the obtained average.
Gomes et al. ( 2012) reported a FE of 1.63, while Mendes et al. (2010) observed an average FEc of 1.26 for Saanen and Brown-Alpine goats, respectively, receiving corn silage.When roughage sources as the corn silage, that present a lower cell wall content and grains, are added to the ruminant diets, better conditions of rumen fermentation are promoted, which contributes to greater feed efficiency values, when compared to those results obtained with dry roughage such as the used oat hay.
The production of fat, protein, lactose, and total solids of the milk (g/d) were not influenced (P>0.05) by the inclusion of CSFA in diets, although with regards to the concentration of these components, there was a negative linear effect (P<0.05) on protein content, with a decrease of 0.87 g/kg milk for each 1% CSFA included in the diet (Table 3).
One of the possible explanations for the linear reduction in the observed concentrations of milk protein, according to the inclusion of protected fat in diets, can be related to the reduction in rumen fermentable organic matter; this effect may be caused by the replacement of  (Geron et al., 2011).This process reduces microbial protein synthesis in the rumen, which is the main source of metabolizable protein that is needed for milk protein synthesis (Santos;Pedroso, 2011).Moreover, obtaining an optimal proportion between energy and protein is important, as it can be used to obtain better results for microbial production, better use of nutrients from the diet, and can consequently improve performance (Rodrigues et al., 2007).
The number of somatic cells was not influenced with the addition of CSFA in the diets, and the obtained average (3.32 log 10 cells/mL × 1000) is close to values of 3.26 log 10 cells/mL × 1000 reported by Ribeiro et al. (2008) in goats.However, Gomes et al. (2012) observed values of 2.84 and 3.07 log 10 cells/mL × 1000 in post peak periods and at the end of lactation, respectively, for multiparous Saanen goats.Although there is currently no legal limit of the number of somatic cell count for goat milk in Brazil (Brasil, 2000) the values observed are higher than the limit of 1,000,000 cells/mL (3.0 log 10 cell/mL × 1000) established by Food and Drug Administration in the United States of America.
The inclusion of CSFA in diets of goats did not influence the concentrations of urea in the serum or in the milk (P>0.05),thus indicating a satisfactory metabolic status; in other words, an adequate amount of nitrogen and energy was provided by the diets (Table 4).Ribeiro et al. (2008), after providing alfalfa hay and corn silage as roughage sources to dairy goats, reported a blood urea concentration of 94.22 and 117.22 mg/dL, respectively, but when the oat hay was the roughage source, the average corresponded to 74.22 mg/dL blood urea, a value that was close to the observed value (61.97 mg/dL).Inclusion of protected fat... 169 Ciênc. Agrotec., Lavras, v.39, n.2, p.164-172, mar./abr., 2015 The concentration of milk urea can be considered a good indicator of metabolism and protein intake for dairy animals, and can be utilized for dietary evaluation (Fernandes et al., 2008).However, the high positive linear correlation (r = 0.9288) that was observed between the results of urea in the milk and blood (Mouro et al., 2002) allows for the choice of one of these methods to evaluate protein animal metabolism.
According to Maia et al. (2010), changes in the blood urea concentration are related to the rumen ammonia content, which is transformed by rumen microorganisms to microbial protein, the main milk protein source.If the available energy for these processes in the rumen is insufficient, ammonia accumulation and an increase in the blood urea concentration can occur, which was not observed with the inclusion of protected fat in diets (Table 4).
The inclusion of CSFA in the diets did not influence (P>0.05)milk fat content; however, the concentration of fatty acids was modified.There was a linear reduction (P<0.05) in the concentration of stearic acid (18:0) and a quadratic effect (P<0.05) on the concentration of linolenic acid (18:3 n-3) (Table 5).
With regards to the degree of saturation of fatty acids identified in milk fat, the concentrations presented  in decreasing order as follows: saturated (65 g/100 g) > monounsaturated (25.22 g/100 g) > polyunsaturated (9.77 g/100 g).These results were more interesting than those obtained by Queiroga et al. (2007) for goat milk; their average saturated, monounsaturated, and polyunsaturated acid values were 75.91, 21.89 and 2.19 g/100 g fatty acids, respectively, due to the greater unsaturated fatty acid content.Among the saturated fatty acids, those that are highlighted in goat milk are capric (10:0), myristic (14:0), palmitic (16:0), and stearic (18:0), which were observed in greater concentrations (Costa et al., 2008), and corresponded to the obtained values (Table 5).
The reduced concentration of stearic acid (18:0) is related to the activity of the ∆9 -desaturase enzyme in the mammary gland that catalyzes the insertion of a double bond in its molecule, converting C18:0 to oleic acid (18:1); however, a difference was not observed (P>0.05) in the concentration of oleic acid in the milk fat.
The quadratic effect (P<0.05) observed with the incorporation of linolenic acid (18:3 n-3) in the milk fat favored a concentration with a maximum point of 24 g CSFA per kg dry matter.Although, when Titi (2011) supplemented goats from the beginning until the intermediate phase of lactation, they reported an increase in the concentration of linolenic acid (18:3 n-3) with the supplementation of 50 g protected fat per kg dry matter.
When evaluating the effects of increasing levels of energy in the diets of goats with the addition of CSFA (0, 28.7, 54.6 and 80.5 g/kg DM), Souza et al. (2014) observed that supplementation improved milk yield with higher levels of essential fatty acids omega-3 (n-3) and omega-6 (n-6).
Although a quadratic effect (P<0.05) was seen in the concentration of 18:3 n-3 (Table 5) the supplementation of CSFA did not increase the total concentration of omega-3 fatty acids, and therefore, did not alter the omega-6/omega-3 ratio.

CONCLUSION
Calcium salts of fatty acids can be included in the diets of goats without interfering with intake and milk production, with improvement of the concentration of linolenic acid (18:3 n-3); nevertheless, the concentration of milk protein is reduced.

Table 1 :
Chemical composition of the experimental diets.

Table 3 :
Milk production and composition of Saanen goats receiving diets with protected fat.

Table 4 :
Urea concentrations in blood and milk of Saanen goats receiving diets with protected fat.

Table 5 :
Fatty acids concentration (g/100g fatty acids) of fat milk of Saanen goats receiving protected fat in diets.