It has been recognized that components of foods can be contributing factors in human health and disease prevention. Based on the potential benefits to long-term human health there is interest in developing sustainable nutritional strategies for reducing saturated and increasing specific unsaturated fatty acids in ruminant milk. Despite the lower scale of milk production from goats compared with cows in Europe, there is an increasing interest in goat milk due to inherent species-specific biochemical properties that contribute to nutritional quality. Goat milk has been identified as a viable alternative for consumers that are sensitive or develop allergic reactions to bovine milk.
1.1. Synthesis and composition of goat milk fat
Fat composition in goat milk is one of the most important components of the technological, nutritional or dietetic quality of goat milk. Milk fat content in goat milk is high after parturition and then decreases during the major part of lactation. This is related to at least two phenomena: a dilution effect due to the increase in milk volume until the lactation peak, and a decrease in fat mobilization that decreases the availability of plasma non-esterified fatty acids, especially C18:0 and C18:1, for mammary lipid synthesis (Chilliard et al., 2003). Even that, total solids, fat, crude protein, lactose, and ash contents of goat milk are almost similar to cow milk, there are important differences in the individual fatty acids and casein fractions and fat globule sizes. Fat globules of goat milk are smaller in size and do not coalesce upon cooling because of lack of agglutinin, which is responsible for the aggregation of fat globules in cow milk.
Goat milk fat is composed primarily of triglycerids (or triacylglycerides) (in 98%) and in a small part from phospholipids and sterols. Triglycerids are synthesized on the outer surface of the smooth endoplasmic reticulum of the milk alveolar cells from precursor substances: fatty acids and glycerol. They are forming larger globules, which are travelling to the margin of cell. At the beginning, they attach to the membrane and they pass through. Then, they are eliminated from the cell as fat globules of the milk. The synthesis is endogenous in a large extent, where the presence of the conjugated linoleic acid plays an important role (Hurley, 2009).
Fatty acids in goat milk are synthesized in epithelial cells of the mammary gland de novo or they are passing over from the blood (Chilliard et al., 2003). Two coenzymes have a major role in the synthesis of fatty acids in goat milk: acetyl-coenzyme A-carboxylase, which participates in the synthesis of fatty acids de novo and fatty acid synthase, which is a complex of enzymatic active substances and is responsible for the extension (elongation) of the fatty acid chain (Hurley, 2009). Fatty acids of exogenous origin are presented via the circulation to mammary epithelial cells either in the form of non-esterified fatty acids or esterified as the acyl groups of the triacylglycerol component of lipoprotein particles. In the mammary gland of ruminant animals, short and medium chain saturated fatty acids are the major products of de novo lipogenesis whereas plasma lipids contribute longer chain and mono unsaturated species. The acetate is the precursor of fatty acids synthesis in ruminants, while in monogastric animals, the precursor is glucose (Clegg et al., 2001).
1.2. Fatty acid composition in goat milk fat
Average goat milk fat differs in contents of its fatty acids significantly from average cow milk fat, being much higher in butyric (C4:0), caproic (C6:0), caprylic (C8:0), capric (C10:0), lauric (C12:0), myristic (C14:0), palmitic (C16:0), linoleic (C18:2), but lower in stearic (C18:0), and oleic acid (C18:1) (Table 1). Three of the medium chain fatty acids (caproic, caprilyc, and capric) have actually been named after goats, due to their predominance in goat milk. They contribute to 15% of the total fatty acid content in goat milk in comparison to 5% in cow milk (Haenlein, 1993). The presence of relatively high levels of medium chain fatty acids (C6:0 to C10:0) in goat milk fat could be responsible for its inferior flavour (Skjevdal, 1979).
|Fatty acid||Goat milk1||Goat milk (from highland flock)2||Goat milk (from mountain flock)2||Cow milk1|
1.3. The effect of nutrition on goat milk fat and fatty acids composition
Nutrition (forage-to-concentrate ratio, type of forages, etc.) is the main environmental factor regulating milk fat synthesis and fatty acid composition in ruminants (Nudda et al., 2003; Bernard et al., 2009). Forage in the diet is known to affect milk fat composition responses to plant oils, including trans-18:1 and conjugated linoleic acid isomer concentrations. Inclusion of fat in the diet enhances milk fat secretion in the goat in the absence of systematic changes in milk yield and protein content (Bernard et al., 2009; Chilliard et al., 2003, 2007). Bernard et al. (2009) found out that, changes in goat milk fatty acid composition were dependent on forage type and plant oil composition, with evidence of an interaction between these nutritional factors. Responses to lipid supplements were characterised as a reduction in fatty acids synthesised de novo (C10:0–C16:0) and an increase in C18:0, cis-C18:1, conjugated linoleic acid and polyunsaturated fatty acid concentrations, indicating that plant oils can be used to effect potentially beneficial changes in milk fat composition without inducing detrimental effects on animal performance. Moreover, goats fed a high level of pasture forage had higher milk fat contents of C4:0, C6:0, C18:0, C18:l, C18:3, C20:0, iso-, ante-iso-, and odd fatty acids, but lower values of C10:0, C12:0, C14:0, C16:0, and C18:2, than those fed the low levels of forage. However, high levels of alfalfa forage also produced the lowest contents of the less desirable trans-C18:1 fatty acids (LeDoux et al., 2002). The conclusion was that decreasing the fibre content and increasing the grain part in the goat daily ration would lead to higher contents of the undesirable trans-C18:1 fatty acids in milk. The composition of goat milk fatty acids differed also in goats grazing one flock on highland (615-630 m altitude) and one flock on mountain (1060-1075 m altitude) pasture by Žan et al. (2005). The most abundant fatty acids in milk of both flocks were C16:0, C18:1, n−9, C14:0 and C10:0 (Table 1). The average content of saturated fatty acids was 74.52 and 73.05% in milk from the highland and mountain flocks, respectively. Three saturated fatty acids (caprylic (C8:0), capric (C10:0) and lauric acid (C12:0)), were present at significantly higher amounts in milk from the highland flock than in milk from the mountain flock. Monounsaturated fatty acids represented 20.49 and 22.32% and polyunsaturated fatty acids 3.73 and 3.24% of the milk from the highland and mountain flocks, respectively. Among the monounsaturated fatty acids, palmitoleic + palmitelaidic acid (C16:1, n−7) showed a significantly higher concentration in milk from mountain flock than in milk from the highland flock. The content of linolelaidic acid (C18:2, n−6) was significantly higher in comparison to milk from the highland flock. The average quantity (32 mg 100 g−1 milk) of essential α-linolenic acid (C18:3, n−3) was slightly higher in milk of the highland flock than in milk from the mountain flock (26 mg 100 g−1 milk). Hou et al. (2011) stated that the supplementation of fish oil can significantly increase the production of cis-9, trans-11 conjugated linoleic acid, and trans-11 C18:1, while lowering the amount of trans-10 C18:1 and trans-10, cis-12 conjugated linoleic acid in the ruminal fluid of goats. Increased cis9, trans-11 conjugated linoleic acid, and trans-11 C18:1 can lead to a higher output of cis-9, trans-11 conjugated linoleic acid in milk product, and the decrease in trans-10 C18:1 and trans-10, cis-12 conjugated linoleic acid supports the role of fish oil in the alleviation of milk fat depression.
1.4. Conjugated linoleic acid
Conjugated linoleic acid consists of a series of positional and geometric dienoic isomers of linoleic acid that occurs naturally in foods. It is a product of biohydrogenation in the rumen of ruminants and has a great influence on synthesis of fatty acids in milk in low concentrations (Bessa et al. 2000; Chouinard et al. 1999; Griinari & Bauman, 1999; Griinari et al. 2000; Khanal & Dhiman, 2004). Actually, the conjugated linoleic acid found in goat milk fat originate from two sources (Griinari & Bauman, 1999). One source is conjugated linoleic acid formed during ruminal biohydrogenation of linoleic acid (C18:2 n-6) that leads first to vaccenic (trans-11 C18:1) and finally to stearic acid (C18:0) (Nudda et al., 2003). The second source is conjugated linoleic acid synthesized by the animal’s tissues from trans-11 C18:1, another intermediate in the rumen biohydrogenation of unsaturated FA. Thus, the uniqueness of conjugated linoleic acid in food products derived from ruminants relates to the incomplete biohydrogenation of dietary unsaturated fatty acids in the rumen. Ruminal biohydrogenation combined with mammary lipogenic and ∆-9 desaturation pathways considerably modifies the profile of dietary fatty acids and thus milk composition (Chilliard et al., 2007).
Dietary sources from ruminants such as milk, cheese and meats contain more conjugated linoleic acid than foods of non-ruminant origin (Bessa et al. 2000; Khanal & Dhiman, 2004). The increase of linoleic acid intake is one of the feeding strategies for conjugated linoleic acid enrichment in ruminant fat since linoleic acid is the main precursor of conjugated linoleic acid (Bessa et al., 2000). The main available sources of linoleic acid in animal feeds are cereal and oilseed grains or oils obtained from these. Goat milk conjugated linoleic acid content increases sharply after either vegetable oil supplementation (Bernard et al., 2009) or fresh grass feeding containing unsaturated fatty acids, but does not change markedly when goats receive whole untreated oilseeds (Chilliard et al., 2003). Mir et al. (1999) found that it is possible to increase conjugated linoleic acid content of goat milk by manipulation of dietary regimen such as supplementation with canola oil. The pasture has major effects by decreasing saturated fatty acids and increasing fatty acids considered as favourable for human health (C9-18:1, C18:3n-3 and C9t11-CLA), compared to winter diets, especially those based on maize silage and concentrates (Chilliard et al., 2007). Investigations have shown that milk fat conjugated linoleic acid content can be also enhanced by manipulation of the rumen fermentation (Bessa et al., 2000; Griinari et al., 1999) or by direct addition of a dietary supplement of conjugated linoleic acid (Lock et al., 2008).
1.5. Effect of fatty acids on health
Milk, apart from its nutritional traits, contains substances which have beneficial effects on human health and is, therefore, considered essential to a correct nutrition. In particular, in milk are present vitamin A, vitamin E, β-carotene, sphingomyelins, butyric acid, and conjugated linoleic acid, all with a strong antitumor effect (Parodi, 1999). Different FA (short and medium chain, saturated, branched, mono and polyunsaturated,
The physiological and biochemical facts of the unique qualities of goat milk are just barely known and little exploited, especially not the high levels in goat milk of short and medium chain fatty acids, which have recognized medical values for many disorders and diseases of people (Haenlein, 2004). Goat milk exceeds cow and sheep milk in monounsaturated, polyunsaturated fatty acids, and medium chain triglycerides, which all are known to be beneficial for human health, especially for cardiovascular conditions. Capric, caprylic acids and medium chain triglycerides have become established medical treatments for an array of clinical disorders, including malabsorption syndromes, chyluria, steatorrhea, hyperlipoproteinemia, intestinal resection, premature infant feeding, non-thriftiness of children, infant malnutrition, epilepsy, cystic fibrosis, coronary by-pass, and gallstones, because of their unique metabolic ability to provide direct energy instead of being deposited in adipose tissues, and because of their actions of lowering serum cholesterol, inhibiting and limiting cholesterol deposition (Alferez et al., 2001; Greenberger & Skillman, 1969; Kalser, 1971; Schwabe et al., 1964; Tantibhedhyanangkul & Hashim, 1978).
Conjugated linoleic acid was recognized as having antioxidative and anticarcinogenic properties in animal model studies (Ip et al., 1991; Jiang et al., 1996; Parodi, 1997). Several
Somatic cells in milk are the total sum of white blood cells present in milk and udder epithelial cells, which may be an indicator of the udder health status (Das & Singh, 2000; Manlongat et al., 1998; Zeng & Escobar, 1996; Wilson et al., 1995). They are present in milk all the time. In cows, a somatic cell count above the regulatory standard is generally considered as an indication of mastitis. An increased number of somatic cell count is either the consequence of an inflammatory process due to the presence of an intramammary infection or under non-pathological conditions due to physiological processes such as oestrus or advanced stage of lactation. For this reason, the somatic cell count of milk represents a sensitive marker of the health of the udder and is considered a useful parameter to evaluate the relationship between intramammary infection and changes in milk characteristics. The standard for the permissible number of somatic cell count for cow milk exists, while it is still under study for goat milk due to considerable fluctuations. When the udder is tired during late lactation, the number of somatic cells in normal conditions can considerably enlarge, and approximately 80% of the cells may be polymorphonuclear leukocytes (Manlongat et al., 1998). The same authors found that normal nonmastitic late-lactation-stage goat milk is significantly higher in polymorphonuclear leukocytes chemotactic activity than early-lactation-stage goat milk. The chemotactic factor(s) present in the milk of normal late-lactation-stage goats is nonpathological and may play a physiologic regulatory role in mammary gland involution. On the other hand, the increase of leucocytes is a response to the inflammatory process in the mammary gland or somewhere in the body. The number of leucocytes increases due to bacterial infections, but it could also be increased due to the stage of lactation, age of the animal, stress, season of the year, nutrition and udder injuries. The variability of somatic cell count in goat milk is very high, which exists among the animals and within the time span of individual animals (Das & Singh, 2000). Therefore, it is important to determine how nutrition can influence the reduction of somatic cell count in goat milk. Gantner & Kompan (2009) found that a five-day supplementation of α-linoleic acid in Alpine goat diet had a significant effect on lower somatic cell count in milk. Based on this experiment, it was concluded that α-linoleic acid supplementation had no effect on milk yield; it had low effect on milk components and significant effect on somatic cell count. A decrease in somatic cell count was determined in the 1st day of the treatment period and continued until 30th day after the treatment period. The supplementation of the goat diet with α-linoleic acid could be used as a method of choice for reduction of somatic cell count in goat milk.
The aim of our study was therefore to ascertain the changes in goat milk yield and its contents of fat, protein, lactose, dry matter, somatic cell count, and total number of microorganisms when goats are supplemented with the following fatty acids: α-linoleic acid, eicosapentanoic acid, and docosahexanoic acid and how these three fatty acids influence on the content of particular fatty acids during and after the supplementation.
2. Material and methods
The research was performed on the farm with 90 Slovenian Alpine and Slovenian Saanen goats. Goats were machine milked. During the experiment, goats were in different stages of lactation. The average body weight of the goats was 51 ± 6 kg. All kids were weaned. Goats were arranged into three pens according to their stage of lactation, namely, after kidding from the forth to the tenth week of lactation (pen A), from the 11th to the 20th week of lactation (pen B), and after the 20th week of lactation (pen C). Goats were milked twice a day, at 6 a.m. (± 30 min) and at 6 p.m. (± 30 min). Diet was composed from hay (2 kg/animal/day) which was given to goats twice a day. Goats were supplemented with feed mixture at milking parlor during the milking time. Supplemental feed mixture contained 50% of grounded maize grains, 30% of dried beet pulp, and 20% of wheat bran. Goats from pen A were supplemented with 500 g, goats from pen B with 350 g, and goats from pen C with 250 g of feed mixture. Vitamin-mineral supplement and water were offered to goats
2.2.1. Measuring performance and milk sampling
The whole experiment was performed in three periods:
2.2.2. Milk yield measuring
There were 90 goats all together in the flock, which were milked on the milking parlor with 24 places for milking goats connected to milk pipeline. Goats were milked every morning between 5:40 and 7:20 a.m. and every evening between 6:20 and 8:00 p.m. A measuring gauged flask was connected to milking unit to measure milk yield. Milk yield was written down for every goat. A milk sample was also taken for the analysis. During the experiment, 30 daily records were collected for every goat, which means 60 records for each goat and 60 milk samples by 70 ml for milk analysis (sample A) and 60 samples by 2 ml (sample B) for fatty acid analysis. The preservative azidiol on the basis of NaN3 in concentration 0.02% with the addition of chloramphenicol for the stabilization of microorganisms was added to the sample A. For every 50 to 70 ml of the milk sample, 0.2 ml of the preservative was added. Milk samples A were then delivered to the Laboratory for dairying, while milk samples B were delivered to the Chemical laboratory at Biotechnical Faculty in Ljubljana.
2.2.3. Analyses of milk samples
2.2.2. Statistical analysis of the data
The statistical package SAS (SAS/STAT, 2000) and partly the statistical package S-PLUS (1966) were used to analyse the data. The statistical analysis did not include records collected during the first six days of the preparation period. In the meantime, the situation in the stable was stabilizing and the team who participated in the experiment was introducing in the everyday milk measuring and collecting samples.
Due to the large fluctuations in individual values of the somatic cell count and number of microorganisms among animals and among observations within animals, we analyzed each animal individually as its time series, and for the most variable ones the logarithm of the values was found (X = log10Y).
The time series were first standardized (S) in the way that last four days (from the7th to 10th day) of the preparatory period (before supplementing with fatty acids) were took as a starting point. Mean value of this period was calculated by the median (Me), the measure of variability was the average absolute deviation (AD). In this way we reduced the impact of outliers. Although, it is usual to standardize by the average and standard deviation, we decided for median and absolute deviation. In this way, the standardized time series for the animal was calculated using the following equation:
In this way, the standardized time series (S) are comparable for animals with different values. Then, we calculated the median for the three periods on the standardized time series:
median for the period from the seventh to tenth day of the experiment (preparatory period), which was in all cases zero (=0);
median for the period from 11th to the 15th day of the experiment (the period of supplementing with fatty acids);
median for the period from the 16th to the 63rd day of the experiment (the post supplemention period of the fatty acids).
For each animal, the corresponding median has become an input data for the statistical analysis. In this way, we analyzed milk yield (ml), the content of milk proteins (g/100 ml), milk fat (g/100 ml), milk lactose (g/100 ml), dry matter (g/100 ml), non-fat dry matter (g/100 ml), total number of microorganisms (n*103/ml), and somatic cell count (n*103/ml) in milk.
In this way, a comparison of groups with a simple analysis of variance was made where the zero assumption was checked for that the averages by groups were the same. If a statistically significant difference test was found (5% level of significance was considered), then the groups were compared also by the Duncan test or by the contrast analysis, where each group was compared with the control group.
All other traits were analyzed by the GLM procedure (General Linear Model) with statistical package SAS, which included the impacts of the group (4) and period (3). Differences among groups were estimated by the linear contrasts, while connections between the properties were calculated by the Pearson correlation coefficient. The limit of statistical significance was taken at P <0.05 and highly statistically significance was taken at P <0.001.
3. Results and discussion
3.1. Milk yield and the chemical composition of milk
The average milk yield and its content of fat, proteins, lactose, dry matter, non-fat dry matter, total number of microorganisms, and somatic cell count in different periods of the experiment by groups is shown in Table 2. In the preparatory period, only somatic cell count statistically significantly differed among groups. Statistically significant differences among groups in the experimental period appeared in dry matter, somatic cell count, and logarithm of the somatic cell count. In the third period of the experiment, statistically significant differences among groups appeared in the majority of observed traits.
It seems that the short time fatty acid supplementation into goat’s diet does not negatively affect their milk yield. Milk yield did not vary statistically significant during the observed period (Table2). As found by Sampelayo et al. (2002), the supplemented fatty acids into the diet of Granadina goats did not affect their milk yield and the content of fat, proteins, lactose, and dry matter in milk.
|Fat (g/100 ml)||3.05a||3.15a||3.00a||2.99a||2.65a||3.40b||2.52a||2.91a||2.84a||3.30b||2.77a||3.06a|
|Proteins (g/100 ml)||2.93a||3.12a||2.98a||3.01a||3.01a||3.21b||3.01a||3.06a||3.15a||3.28a||3.29a||3.07b|
|Lactose (g/100 ml)||4.59a||4.55a||4.49a||4.53a||4.58a||4.54a||4.48a||4.44a||4.50a||4.54a||4.44a||4.46a|
|NFDM (g/100 ml)||8.32a||8.48a||8.26a||8.33a||8.39a||8.55a||8.29a||8.30a||8.45a||8.62b||8.53a||8.33a|
|DM (g/100 ml)||11.37a||11.62a||11.26a||11.33a||11.04a||11.76b||10.81a||11.21a||11.29a||11.92b||11.30a||11.39a|
Milk fat yield statistically significantly increased in ALFA group from 3.15 to 3.40 g/100 ml on average when goats were supplemented with linseed oil rich in α-linoleic acid (Table 2) and it slightly decreased to 3.30 g/100 ml until the third period of the experiment. In groups EPA and DHA, milk fat yield firstly decreased, while it increased slightly after the end of supplementation with fatty acids.
There were no statistical significant differences among the groups of goats in milk protein yield before the supplementation with fatty acids (Table 2). During the supplementation of goats with fatty acids, milk protein yield increased and it was increasing also after the end of supplementation. Group ALFA had the highest protein yield in milk in the whole time of the experiment.
In general, lactose in milk varies little, what was confirmed also in our research. There were no statistical significant differences in lactose yield among the observed groups, neither during the supplementation with fatty acids nor after that (Table 2).
Non-fat dry matter increased during the experiment in all observed groups which were supplemented with fatty acids, but not in the control group KONT (Table 2). Differences among groups were not statistically significant. Total dry matter decreased after supplementing with fatty acids in groups EPA, DHA, and KONT, while it increased in ALFA group. After the end of supplementing with fatty acids, total dry matter increased in all groups. Group ALFA statistically significantly differed in milk dry matter from other observed groups in the second and third period of the experiment.
The number of microorganisms in milk mostly depends on milking hygiene, which includes staff, animals, facilities, equipment, hygiene maintenance, and cleaning of the equipment. It also depends on the health of the udder and the presence of mastitis. Soon after the beginning of the experiment, the hygiene and cleaning improved, and the number of microorganisms in milk decreased (Table 2). There was no mastitis detected in the whole time of the experiment. No statistically significant differences were noticed among groups in the number of microorganisms in milk.
Somatic cell count was one of the most variable traits in our experiment, since we found that values ranged from 13.000 to 24,312.000 of somatic cells in ml of milk. Despite the great variability, transformation of somatic cell count to the logarithmic value enabled to find the possible impacts of supplementation with fatty acids on somatic cell count (Figure 1). Preliminary report by Košmelj et al. (2001) showed the impact of supplementing alpha-linolenic fatty acid to goats, which was reflected in a reduction of the number of somatic cells during the supplementation and four weeks after.
The average values for medians during the supplementation with fatty acids (Me1) and for medians five days after the supplementation with fatty acids (Me2) are shown in Table 3. Results showed statistical significant differences among groups of goats for medians during the supplementation with fatty acids and also for medians five days after the supplementation with fatty acids. The average of medians (Me1 and Me2) in group ALFA is negative, so it could be affirmed, that the supplementation of linseed oil rich in α-linoleic acid decreases the number of somatic cell count in milk.
|Period / Group||EPA||ALFA||DHA||KONT|
|Average for Me1||1.01b||-3.11 a||1.68 b||3.52 b|
|Average for Me2||1.75 b||-2.47 a||1.78 b||3.20 b|
On average, somatic cells in goat milk are present in a greater number than in cow milk. Zeng et al. (1997) reported that 17% of goat milk samples recorded on goat farms which are members of the Association of goat farmers in the U.S. exceeded the standard 1.0x106 of somatic cells ml-l when the experiment of daily monitoring of somatic cells in milk was carried out.
Das & Singh (2000) studied somatic cells in goat milk and electrical conductivity of milk. In the blood samples total leucocytes and differential leucocytes (lymphocytes, monocytes, neutrophils, eosinophil, and basophils) were also determined. Somatic cell count in goat milk was high during early lactation and decreased subsequently as the lactation advanced. There were found individual variations (P<0.01) in somatic cell counts between different lactation periods as weel as among and within animals. For example, one goat had very high somatic cell count in comparison to other goats from the beginning to the end of the experiment. The goat was then tested for mastitis using California mastitis test and it was found to have normal milk. Similar results were found in our experiment. Total leucocyte count in blood also decreased as the lactation progressed and remained fluctuated during late lactation in the study by Das & Singh (2000). Lymphocytes and neutrophils were low during early lactation and with establishment of lactation stabilized to normal levels. Protein content of milk did not vary during different periods of lactation. However, lactose decreased and fat percent increased with advanced lactation. It is interesting that the connection between somatic cell count and milk yield and between somatic cell count and milk composition was not found in any stage of lactation.
Mastitis is typically associated with a large number of somatic cells in small ruminants. In our experiment, the number of somatic cells significantly reduced only in the ALFA group and lasted statistically significant 39 days after the supplementation with fatty acids. For α-linolenic fatty acid is known, that it could incorporate into phospholipids five hours after ingestion (Adam et al., 1986). The other two, eicosapentaenoic acid and docosahexaenoic acid can incorporate into phospholipids only after a few days supplementation. The statistically significant effect of the α-linolenic fatty acid only on somatic cell count could be explained by the rapidness of incorporation into membrane phospholipids of this fatty acid.
The fluctuations of the somatic cell count in goat milk are subjected to many influences. Researchers have not explored other reasons for the number of somatic cells in goat milk except the hygiene measures. Ruminants are in the last 20 years fed adding n-3 fatty acids to improve the fatty acid composition of milk and meat, but the impact on the number of somatic cells have not been monitored. Our experiment clearly shows that the supplementation of the α-linolenic fatty acid had a relatively long time impact on reducing somatic cell count or to a low level of somatic cells in milk. The interpretation may be possible, that we achieved a more appropriate relationship between n-3 and n-6 long chain fatty acids with the supplementation of α-linolenic fatty acid which was not provided by the diet.
3.2. Composition of fatty acids in goat milk
Chemical analysis of goat milk fat was done for fatty acids from 10:00 to 24:6, n-9. The fat composition of goat milk was studied by each milking during the experiment time. Therefore, values listed below (Table 3) represent the percentage of the all analyzed fatty acids rather than total fat in goat milk.
During our experiment, there was from 9.0 to 14.0 wt % of the capric acid (10:0) in the goat milk fat. Some authors (Hurley, 2009; Jandal, 1996; Sanz Sampelayo et al., 2002) indicated values from 8.4 to 11.1%. EPA group had the lowest level of capric acid before supplementing with fatty acids, while its level exceeded groups ALFA and KONT during the supplementation and declined to the lowest level among groups in the last period of the experiment. DHA group had the highest level of the capric acid during the supplementation with fatty acids as well as all the time after the supplementation. It is known that goat milk has more short-chain fatty acids (C4:0 to C10:0) than cow’s milk, which are easier to digest than long-chain fatty acids.
We found that the
|FA / GROUP||EPA||ALFA||DHA||KONT||EPA||ALFA||DHA||KONT||EPA||ALFA||DHA||KONT|
|18:1, n-9c, 18:1,|
n-9t, 18:1, n-12t, 18:1, n-7c
|20:3, n-3||0.03a||0.02a||0.02a||0.02a||0.32b a||0.03a||0.04a||0.03a||0.12b||0.04a||0.02a||0.03a|
|n-3||0.99 a||0.81 a||0.76 a||0.73 a||4.13 b||3.34 b||1.68 a||1.11 a||1.99 b||1.38 a||1.38 a||1.10 a|
|n-6||2.70 a||2.50 a||2.56 a||2.47 a||3.47 b||3.70 b||3.17 b||2.67 a||2.69 b||2.93 a||2.92 a||2.95 a|
|n-3/n-6||0.37 a||0.32 a||0.30 a||0.30 a||1.19 a||0.90 b||0.53 b||0.42 a||0.74 b||0.47 a||0.47 a||0.37 a|
|n-3 : n-6 (1:X)||2.73 a||3.09 a||3.37 a||3.38 a||0.84 b||1.11 b||1.89 a||2.41 a||1.35 b||2.12 a||2.12 a||2.68 a|
|LC PUFA n-3||0.36 a||0.30 a||0.28 a||0.27 a||3.29 b||0.52 a||3.35 b||0.44 a||1.27 b||0.54 a||1.51 b||0.42 a|
|LC PUFA n-6||0.34 a||0.32 a||0.31 a||0.30 a||0.52 b||0.31 a||0.63 b||0.31 a||0.39 a||0.33 a||0.44 a||0.36 a|
n-3/ LC PUFA n-6
|1.06 a||0.94 a||0.90 a||0.90 a||6.33 b||1.68 a||5.32 b||1.42 a||3.26 b||1.64 a||3.43 b||1.17 a|
|LC n-3 : n-6 (1:X)||3.11 a||2.93 a||2.91 a||3.00 a||12.17 b||5.41 a||8.44 b||4.58 a||8.35 b||4.96 a||7.80 b||3.24 a|
There was between 20 and 29 wt % of the
In goat milk fat, between 1.06 and 1.73 wt % of the
The content of oleic fatty acid (18:1, n-9) was in our experiment determined in the concentration from 19.0 to 28.0 wt %. During the supplementation with fatty acids, the content of oleic acid statistically significantly declined in EPA and DHA groups. An increase of the content of oleic acid in milk was observed in groups KONT and ALFA, as during as well as after the supplementation with fatty acids, but differences in these two groups before and after the supplementation were not statistically significant. Sanz Sampelayo et al. (2002) noted the content of oleic acid in goat milk around 22 to 24% and stated that despite the addition of various concentrations of protected polyunsaturated fatty acids the content of oleic fatty acid in goat milk remained fairly constant.
Conjugated linoleic acid is an intermediate product of the biohydrogenation, therefore its high concentration in DHA group was logical, since the degradation of the docosahexaenoic acid in the rumen is the slowest. The concentration of the conjugated linoleic acid in goat milk fat was relatively high also in ALFA group, knowing that the biohydrogenation of the α-linoleic acid is the fastest (Gulati et al., 1999), what we also observed in an increased concentration of C 18:1 in ALFA group. The conjugated linoleic acid is synthesised in the mammary gland of lactating animals and in the muscles of young animals. In our experiment, the conjugated linoleic acid probably did not originate only from the supplemented fatty acids, what was found also by Griinari et al. (2000).
Before the supplementation with fatty acids, there was from 2.00 to 2.66 wt % of the
There was less than 0.06 wt % of the
The content of
At the beginning of the experiment, the content of
The maximum concentration of
According to the fact that the transfer of eicosapentaenoic acid through diet into the milk can be so effective, it is important how to produce milk enriched with n-3 and n-6 fatty acids. Consumers are increasingly use milk with lower fat content. Thus, milk enriched with n-3 and n-6 fatty acids would significantly help to more correct and balanced diet, especially in children and elderly people.
Before supplementation with fatty acids, the content of
There was from 0.046 to 0.136 wt % of the
Only 0.05 to 0.1 wt % was the concentration of
The effectiveness of transfer the docosahexaenoic fatty acid into milk was observed in cows by Chilliard et al. (2001), which amounted 4.1%. In goats, it amounted 3.5% for unprotected fatty acids and 7.6% for protected fatty acids (Kitessa et al., 2001). The estimated transfer of docosahexaenoic fatty acid in our experiment was 7.84.
There was 53 to 57 wt % of the
As reported Kitessa et al. (2001), a significant decrease appeared in C10 to C16 fatty acids after adding fish oil into the diet for goats, but when Chilliard et al. (2001) fed cows with fish oil only, they noticed a slight decrease in C4 to C14 fatty acids, or even 1.3% increase of these fatty acids when adding fish oil in the duodenum. In the experiment by Kitessa et al. (2001), a group of animals were supplemented a protected fish oil from 19th to 26th day and then unprotected fish oil from the 37th to 42nd day. Due to the significantly reduced feed intake and milk production in sheep the unprotected fish oil was administered a short time. Between one and another type of feeding was only eight days, which is questionable. It is possible that there was an influence of the previous supplementation, because our data showed that the effect of supplementation with some types of fatty acids can take more than 10 days on changes in the fermentation of medium chain fatty acids. Even Sanz Sampelayo et al. (2002) in goats found that the percentage of total unsaturated fatty acids reduced after the supplementation with protected polyunsaturated fatty acids.
The content of
Before the supplementation with fatty acids,
The passage of the supplemented polyunsaturated fatty acids from the gastrointestinal tract into milk was estimated on the basis of the differences between the content of fatty acids before supplementation and the difference between KONT group and other groups during the supplementation and thereafter, taking into account the amount of milked milk during the supplementation and 14 days thereafter. The results are shown in Table 3, where it is clear that the passage of the conjugated linoleic acid into milk was 12.79%, 14.03% of the eicosapentaenoic acid, and 21.13% of the docosahexaenoic acid. The differences were statistically significant (p <0.05).
|Supplemented PUFA during experiment (g)||95||72||75||0|
|PUFA appeared in milk from the 6th to the 34th day (g)||50.78||46.66||53.3||37.45|
|Difference or estimated passage (%)||14.03||12.79||21.13||0|
3.3. Correlations between somatic cell count and some fatty acids
Correlations between somatic cell count and some fatty acids during the experiment were calculated by the Pearson correlation coefficient. The same correlations were calculated also for the second and third period of the experiment (from the 11th to the 65th day) and for the period from the 21st to the 65th day of the experiment. Statistically significant correlations between somatic cell count and C10 throughout the whole experiment were found in EPA group (r=0.24), ALFA (r=-0.18), and (r=-0.17) KONT group. The correlations between somatic cell count and C12 and between somatic cell count and C14 were statistically significant throughout the whole experiment only in EPA group (r=0.25 and r=0.24, respectively; p<0.01). From the 11th to the 65th day of the experiment, there were only correlations between somatic cell count and C10 in DHA group (r=-0.30), between somatic cell count and C12 in DHA group (r=-0.37), and between somatic cell count and C14 in ALFA (r=0.26) and DHA (r=-0.29) groups found statistically significant (p<0.05). From the 21st to the 65th day of the experiment, correlations between somatic cell count and C10 in EPA (r=-0.45) and DHA (r=-0.46) groups, between somatic cell count and C12 in EPA (r=-0.43), DHA (r=-0.53), and KONT (r=0.39) groups, and between somatic cell count and C14 in ALFA (r=-0.59), DHA (r=-0.57), and KONT (r=0.44) groups were statistically significant (p<0.05).
Correlation between somatic cell count and C18:1 was statistically significant only in EPA group (r=-0.24) throughout the whole experiment, in DHA group (r=0.47) from the 11th to the 65th day of the experiment, and in EPA (r=0.42), ALFA (r=-0.49), and DHA (r=0.67) groups from the 21st to the 65th day of the experiment. Between somatic cell count and C18:3, the correlation was statistically significant only in ALFA (r=-0.43) group from the 11th to the 65th day of the experiment. No correlations between somatic cell count and C20:4 throughout the whole experiment were statistically significant. There were only correlations between somatic cell count and C20:4 in EPA group from the 11th to the 65th day of the experiment (r=0.36) and from the 21st to the 65th day of the experiment (r=0.66) statistically significant.
Statistically significant correlation between somatic cell count and monounsaturated fatty acids throughout the whole experiment was found only in ALFA group (r=-0.22) and from the 11th to the 65th day of the experiment in DHA group (r=0.50). From the 21st to the 65th day of the experiment, this correlation was statistically significant in EPA (r=0.43), ALFA (r=-0.50), and DHA (r=0.68) groups. Between somatic cell count and polyunsaturated fatty acids, only the correlation in ALFA group from the 21st to the 65th day of the experiment was found statistically significant (r=-0.49).
Our research proved that the supplementation of fatty acids into the diet had no effect on daily milk yield of goats. In ALFA group, a statistically significant impact on the increase of the protein content in milk (p<0.01) during the supplementation and thereafter was observed. Fat content was increasing during the supplementation and thereafter in ALFA group, while in EPA and DHA groups, fat content significantly reduced during the supplementation with fatty acids (p<0.001) and a few days thereafter. This finding indicates that the supplementation with fatty acids (eicosapentanoic and docosahexanoic fatty acid) had a negative impact on the milk fat production. Lactose content did not change significantly during the supplementation and no differences were found among groups. Non-fat dry matter content was the highest in ALFA group, its increased value reflected even after the end of the supplementation with fatty acids.
The supplementation of α-linoleic fatty acid decreased somatic cell count in milk, even 30 days after the end of the supplementation. Statistically significant decrease of somatic cell count, compared to the period prior to the supplementation, was appeared till the 29th day after the end of the supplementation (p<0.05). The number of micro organisms in milk is the result of hygienic conditions at milking, hygiene of milking personnel, equipment, environment and hygiene of the animals. In the case of our study, it has been established that the lower number of micro organisms was the consequence of better hygiene during the experiment due to the experimentalists’ presence.
The supplementation of α-linoleic, eicosapentanoic and docosahexanoic fatty acids had different effects on the composition of fatty acids in milk fat. Eicosapentanoic fatty acid supplemented into the diet of EPA group increased the following fatty acids: capric, lauric, myristic, conjugated linoleic, linoleic, γ-linolenic, cis-11,14,17-eicosatrienoic, cis-8,11,14-eicosatrienoic, arachidonic, eicosapentaenoic, docosatrienoic, docosatetraenoic, and docosapentaenoic acid. The supplementation of eicosapentanoic fatty acid decreased palmitic, stearic, and oleic fatty acid. α-linoleic fatty acid supplemented to ALFA group increased the following fatty acids: lauric, miristoleic, oleic, conjugated linoleic, linoleic, α-linoleic, γ-linolenic acid. This means that there was no elongation from α-linoleic acid into fatty acids with longer chain. A decrease was observed in myristic, palmitic, and docosatetraenoic acid. DHA group was supplemented with docosahexaenoic fatty acid where the increase of the following fatty acids was recorded: capric, lauric, myristic, palmitoleic, conjugated linoleic, linoleic, γ-linolenic, cis-8,11,14-eicosatrienoic, arachidonic, eicosapentaenoic, docosatrienoic, docosatetraenoic, docosapentaenoic, docosahexaenoic acid, while a decrease was noticed in the following fatty acids: miristoleic, palmitic, stearic, and oleic acid. In the control group, only slight variations in some fatty acid levels were recorded, which were not statistically significant.
Research showed that the supplementation of α-linoleic acid had a positive impact on reduction of the somatic cell count in goat milk. However, the surprising result was found, that the supplementation of eicosapentanoic and docosahexanoic acid did not affect the reduction of somatic cell count in the same extent. There is a question, whether this is the result of the supplement or of the n-3:n-6 ratio which changed after the supplementation. Since the ratio n-3:n-6 changed to the similar value when other fatty acids were supplemented and the effect was not the same, it seems that the n-3:n-6 ratio was not the cause of this effect. It is suggested that α-linoleic acid could be rapidly incorporated into cell membranes, which displace arachidonic acid. This is resulted in more flexible cell membranes and better anti-inflammatory effect. Perhaps this mechanism was the one which contributed to the reduction of somatic cell count. For further research, it would be necessary to also include this kind of analysis. The results also showed that the transition of long chained polyunsaturated fatty acids into goat milk appeared relatively in large extent, therefore, polyunsaturated fatty acids occur in milk fat very quickly after their consumption.