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Fat Profiles of Milk and Butter Obtained from Different Dairy Systems (High and Low Pasture) and Seasons (Spring and Fall): Focus on Healthy Fatty Acids and Technological Properties of Butter

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31 July 2024

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31 July 2024

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Abstract
This study aimed to evaluate the fatty acid (FAs) profile in milk from commercial farms with varying pasture levels in the diet during spring and fall, and to investigate the physical and chemical properties of butter to assess the impact of FAs on technological and nutritional properties. Milk sampling was conducted biweekly from six farms, categorized into high (HP) and low (LP) pasture treatments based on pasture intake: >60% and <35%, respectively. Butter was made from a pasture-based system (GRZ) and a confined system (C). No differences were observed in milk fat percentage between HP and LP in either season. High pasture had 85-66% more conjugated linoleic acid (CLA, p=0.01), 74-48% more trans-vaccenic acid (TVA, p=0.01), and 21-15% more branched- chain FAs (BCFA, p=0.006) than LP in spring and fall, respectively. In fall, butter from C had lower saturated FAs (SFA, p=0.005), higher unsaturated FAs (UFA, p=0.008), and a lower spreadability index (SI, p=0.005) than GRZ, resulting in softer butter. In conclusion, HP in both seasons had higher contents of FAs considered healthy for consumers compared to LP. Contrary to expectations, in fall, C showed higher UFATVA and lower SFA in butter, leading to better technological characteristics than GRZ.
Keywords: 
Subject: Biology and Life Sciences  -   Food Science and Technology

1. Introduction

Milk and dairy products hold significant nutritional value in the human diet, supplying essential energy, protein, vitamins, and minerals [1,2,3,4]. Various factors influence milk composition, including diet, breed, parity, environmental conditions, feeding and management practices, season, and lactation state [5,6]. Among these factors, lipids constitute the most variable fraction of milk and are highly responsive to dietary modifications in terms of composition and concentration [1,3]. The chain length and degree of unsaturation of fatty acids (FAs) are critical determinants of milk fat quality concerning human health. These factors are closely linked to lipid digestion in bovine species. Lipids in the diet undergo lipolysis and biohydrogenation processes mediated by rumen bacteria, leading to the saturation of most consumed unsaturated FAs (UFAs). The progression of this process is influenced by the characteristics (type and quantity) of lipids and the type of diet [1,4,7]. Moreover, it should be noted that some natural fatty materials, such as fat from meat and ruminant milk, contain trans fatty acids, including vaccenic acid (TVA) and conjugated linoleic acid (CLA). Conjugated linoleic acid comprises a family of positional and geometric isomers, all of which are conjugated dienes of linoleic acid (18:2). These FAs have been reported to offer beneficial health effects, including hypocholesterolemic, anticarcinogenic, antiatherogenic properties, modulation of the immune response, and improvement of bone mineralization [8,9,10]. It has been reported that the consumed amount of CLA needed to produce observable health benefits ranges from 0.8 to 3.0 g/d [11]. Vaccenic acid, an intermediate in CLA formation, also possesses beneficial health properties, including the reduction of cardiovascular disease risk and potential inhibition of tumor growth [12]. Additionally, oleic acid and TVA found in milk have also been shown to have beneficial effects on human health, similar to CLA. Oleic acid reduces total cholesterol and low-density lipoprotein (LDL) [13], while TVA is desaturated to CLA in the human body at a rate of 19% [14]. Furthermore, odd- and branched- chain FAs (OBCFAs) are unique components of ruminant milk fat, representing the primary contribution of BFAs to human nutrition. The quantification of milk BFAs has garnered significant interest in recent years. Due to their origin and the correlations observed between diet and ruminal microbial population, OBCFAs are considered potential biological indicators of ruminal function [15]. The potential inhibitory effects on tumor cells [16] and the reduced risk of cardiovascular disease, associated with the consumption of these fatty acids further underscore their beneficial effects on human health, including a lower risk of developing type 2 diabetes [17]. Specifically, 15:0 and 17:0 iso and anteiso have been found to enhance the fluidity of cell membranes [18]. The concentration and composition of milk fat can be readily modified through dietary adjustments [19]. For example, increasing the proportion of forage in the diet compared to concentrate results in higher concentrations of OBCFAs in the milk. Similarly, a diet rich in grass silage could elevate the total content of OBCFAs in milk. The profile of OBCFAs in cow’s milk is primarily influenced by the FAs in the diet and FAs metabolism in the rumen [20]. Therefore, understanding the origin of OBCFAs in milk and manipulating the diet of dairy cows to produce milk enriched with odd- and branched- chain FAs can be important both scientifically and industrially.
Prior studies have investigated milk fatty acids and their correlation with human health using certain indices. The atherogenic index (AI) is indicative of the risk impact on cardiovascular diseases. A higher AI suggests a greater risk of such diseases [21]. Additionally, the ratio between hypocholesterolemic (18:1, 18:2, and 18:3) and hypercholesterolemic FAs (12:0; 14:0, and 16:0), denoted as H/H, is associated with AI [22]. Another index associated with human health is the n-6/n-3 ratio [23]. Excessive levels of n-6, commonly found in Western diets, can hinder human enzymatic systems, contributing to the development of certain diseases. Conversely, higher levels of n-3 can have the opposite effects [23,24]. Furthermore, the technological potential of milk for butter production was assessed using the spreadability index (SI) [25].
Moreover, pasture-based systems are regarded as more environmentally friendly, animal welfare-conscious, and sustainable compared to confinement systems [26]. Additionally, milk and dairy products from these systems offer potential nutritional benefits and market opportunities due to their improved composition compared to those derived from total mixed rations (TMR) systems [27,28]. Pasture-based systems are commonly employed in regions with mild climates like South America and Oceania, owing to their low production costs, which are favored by climate conditions and forage accessibility [29]. However, even in these systems, the use of reserves and concentrates is necessary to ensure the fulfillment of energy requirements and nutrient quality [29,30]. In Uruguay, pasture utilization is crucial for reducing production costs and maintaining satisfactory production levels [29]. Furthermore, it has been widely reported that including pasture improves the FAP in milk and dairy products [31,32]. For instance, milk and dairy products from systems with a high proportion of pasture exhibit higher levels of UFA and CLA, and lower proportions of saturated fatty acids (SFA) compared to systems with low levels or no access to pasture [25,33,34,35,36,37,38]. Therefore, the ability to modify the composition of milk fat through pasture utilization and strategic supplementation can serve as a tool to differentiate dairy products, resulting in milk and dairy products with a healthier FAP for human consumption. It is important to consider that this management approach has its limitations. For instance, the addition of UFA to the diet increases CLA production, but excessive amounts can result in a lower percentage of milk fat by reducing the production of new FAs in the udder [7,39]. Moreover, seasonality, particularly in pasture-based systems, affects the FAP in milk and dairy products. Climate factors influencing the thermal comfort of cows, primarily heat stress in summer, impact the lipid catabolism of the animals [40]. Additionally, seasonality impacts the quality of pastures [41], thus significantly influencing the quality of milk fat. In this regard, higher levels of monounsaturated FAs (MUFAs), PUFAs, and CLA, along with a lower n-6/n-3 ratio and palmitic acid content, have been reported in milk and cheeses in spring compared to fall [42,43,44]. This may result from lower pasture availability and increased utilization of preserved forages in temperate countries during fall, leading to decreased FA quality in the cows’ diet [29,43].
In the past, the consumption of dairy fat (such as milk cream, butter, or cheese) raised concerns among consumers due to its high levels of SFA, which had been linked to elevated cholesterol levels, arteriosclerosis, and cardiovascular diseases [21,45]. However, recent reviews and meta-analyses have concluded that milk consumption has at least a neutral effect on various health outcomes, and cow’s milk consumption may even be beneficial for osteoporosis, cardiovascular disease, stroke, type II diabetes, and certain cancers [46,47,48]. Regarding organoleptic characteristics, it has been reported that modifications in the FAP result in changes in the texture, sensory, and nutritional quality of butter [49]. In this regard, butter from pasture-based systems has shown better nutritional and rheological quality compared to butter from confinement systems using TMR [50]. Additionally, higher levels of polyunsaturated fatty acids (PUFAs), including CLA, and a higher yellow index have been reported in milk from pasture-based systems compared to TMR systems [51].
This study aimed to evaluate the FAP (mainly OBCFA) in milk obtained from farms located in the northwest region of Uruguay, using feeding strategies with varying pasture content (classified as high and low pasture in the diet) during two seasons (spring and fall). The physical and chemical properties of butter were investigated to assess the impact of varying fatty acid profiles on the technological and nutritional properties of high-fat dairy products.

2. Materials and Methods

2.1. Experimental Design, Localization and Samples Analysis

Six dairy farms were selected based on the pasture intake, milk production, and somatic cell count from the previous year. Two treatments were created according to pasture intake: High Pasture (HP) (>65% pasture of total intake) and Low Pasture (LP) (<35% pasture of total diet intake).
The study was conducted during two seasons: spring 2021 and fall 2022. Milk samples were collected from bulk tanks (if there were multiple tanks, a proportional mixture was prepared based on the volumes of each tank), considering that the milk originated from two or four milkings. Sampling was conducted fortnightly. The samples were extracted in 15 mL Falcon tubes, frozen for transfer, and subsequently analyzed in the laboratory of Food Technology at the School of Chemistry, Universidad de la República. Concurrently, feed samples (provided to dairy cows), including pasture and supplements (concentrate and reserves), were collected. In each dairy farm, the following records were kept: herd management, milk production, feeding routine (type of supplement, quantity, and composition), type of pasture (allocation, availability, and species), milk production, and number of milking cows.
During the spring, the HP treatment had an average of 74% pasture and 26% supplement, with a forage/concentrate ratio of 79:21, while the LP treatment had 10% pasture and 90% supplement, with a forage/concentrate ratio of 68:32. In the fall, the HP had an average of 45% pasture and 55% supplement, with a forage/concentrate ratio of 76:24, while the LP had 3% pasture and 97% supplement, with a forage/concentrate ratio of 61:39.
Pastures consisted of a variety of grass species (Festuca arundinacea, Avena sativa) and legumes (Medicago sativa, Lotus corniculatus, Trifolium pretense, Trifolium repens), as well as natural grassland. The ingredients used in commercial dairies varied over time, depending on available conserved forage and the market availability of grains and by-products for concentrate. The most commonly used conserved forages were whole plant maize or sorghum silage, as well as grass hay, silage, or haylage (such as oat, lucerne, and moha). Concentrate mixes could include ground corn grain, rice bran, soybean expeller, soybean meal, barley rootlets, urea, yeast, and minerals.
Daily pasture dry matter (DM) intake (kg DM/cow) was estimated using the energy balance method according to NRC [52] guidelines, as the amount of pasture required to provide the remaining energy needed to meet the cow’s net energy (NE) requirement, not supplied by supplements (reserves and concentrates). Cow NE requirements were estimated as the sum of maintenance and milk production requirements [53]. The average data for neutral detergent fiber (NDF), acid detergent fiber (ADF), and ether extract (EE) of each group (pasture and supplement) are presented in Table 1.
Table 2 shows the fatty acid profiles of pasture and supplement in each treatment (HP and LP) (averaged over the experimental period).

2.1.1. Butter Production

Butter was produced from two contrasting dairy farms based on diet and management: grazing + supplement (GRZ) and confinement cows with total mixed ration (TMR) (C: confined). At the time of milk extraction for butter production, a sample of feed (pasture and supplement) was obtained from each producer in each season. For the C (confined) dairy farm in spring and fall, the total mixed ration consisted of ground corn grain and soybean expeller for concentrate, haylage (Medicago sativa) as conserved forage, urea, and minerals. For the GRZ dairy farm in spring and fall, the supplement comprised corn grain and barley rootlet. Pastures consisted of Avena sativa and Medicago sativa in the fall, and Trifolium pretense and Lolium multiflorum. The FAP in the diet of cows from each farm (GRZ and C) is shown in Table 3.
Two butter batches were produced from each producer. For each butter production, 60 L of bulk milk (MilkB) were obtained, which was immediately transported under refrigerated conditions (4 °C) to the Universidad Tecnológica del Uruguay (UTEC) dairy pilot plant (La Paz, Colonia, Uruguay) for processing. Two batches of fresh cream (5 L each; 40–42% fat) separated at 45 °C from pre-pasteurized milk. The cream was then stored overnight at 10 °C for maturation. Subsequently, both cream samples were churned in a rotary churn (Edibon España) at 26 rpm and 13 °C. After grain formation, the butter was washed with 1.7 L of water at 4 °C, and finally, it was kneaded until it formed into butter. Portions of butter were clarified for further analysis using the British Standards Method 769 (BSI 1961 54) [54].

2.2. Samples Analysis

2.2.1. Fatty Acid Profile in Milk and Butter

Milk and butter samples (3 g each) were extracted using the Rose-Gottlieb technique [55]. Analyses were conducted in triplicate, and fatty acid methyl esters were prepared following the IUPAC 2.301 protocol, [56] and analyzed by gas chromatography according to the AOCS Ce 1c-89 and AOCS Ce 1f-96 protocols, [57] . The gas chromatograph was a Shimadzu (Kyoto, Japan) model 2014 equipped with a Supelco (Bellefonte, PA) SP 2560 (100 m × 0.25 mm × 0.2 mm) capillary column and a flame ionization detector (FID). The injection volume of the samples was 1 μL. The temperature program used was as follows: an initial temperature of 90 °C for 2 minutes, then an increase to 175 °C at a rate of 20 °C/min, maintained for 35 minutes, followed by an increase to 240 °C at a rate of 15 °C/min, maintained for 25 minutes. Peak identification was achieved through the analysis of authentic standards. Standards and reagents used for the analysis were supplied by Sigma–Aldrich (United States). The milk fat compositions were expressed in grams of each individual fatty acid per 100 g of total fatty acids. The atherogenicity index (AI) was calculated as (12:0 + 4 × 14:0 + 16:0) / (MUFA + PUFA) [21], and the hypocholesterolemic/hypercholesterolemic fatty acids (H/H) ratio was calculated as (18:1cis + 18:2cis + 18:3cis) / (12:0 + 14:0 + 16:0) [22] . The spreadability index (SI) was calculated as the ratio of 16:0 to 18:1, as proposed by O’Callaghan et al. [25] .

2.2.2 Fatty Acid Profile in Pasture and Supplement

Fat was extracted from the different samples (1 g each) using the Hara and Radin, [58] technique with a mixture of hexane and isopropanol (3:2). Initially, approximately 1 g of milled samples was weighed into a tube, and 20 mL of the solvent mixture was added. Extraction was conducted at room temperature (20 °C) with magnetic stirring for 90 min. Subsequently, centrifugation was performed (Hermle, model Z 200 A) at 3000 rpm for 15 min. Following centrifugation, 5 mL of the solvent mixture was added for complete lipid extraction. Finally, the total solvent mixture was removed using a nitrogen flow at 40 °C until the weight of the lipids remained constant. Analyses were conducted in triplicate, and the fatty acid profile was determined as previously described for milk and butter.

2.2.3. Butter Hardness

The hardness of butter was measured at 10 °C using a Texture Analyzer (Brookfield CT3 50k, USA) following the procedure [59] with modifications. The procedure consisted of a testing the cutting force at depth of 16 mm applied to a given sample, that was stored at 10°C for 12 h, at a speed of 2 mm/s using a TA53 cutting wire probe. The force of cutting between 8 and 16 mm was reported as the firmness of the sample. Each sample was measured six times.

2.2.4. Differentials Scanning Calorimeter (DSC)

A Shimadzu differential scanning calorimeter (DSC-60A plus, Shimadzu Co., Japan) was used to examine the melting and crystallization properties of milk fat from butter. Oxygen (99.999% purity) was used as the purge gas. The DSC was calibrated using high-purity indium (m.p. 156.6 °C, ∆Hf = 28.45 J g−1) according to standard DSC procedures. Clarified butter samples of about 10 mg were placed in open aluminum pans and inserted into the heating chamber of the DSC cell, with an empty aluminum pan as the reference. Prior to analysis, the samples were heated at 50 °C for 5 minutes to melt all crystals and nuclei. Subsequently, they were tempered in a freezer at -20 °C for 48 hours. The final crystallization was carried out in the DSC by cooling from 2 °C to -50 °C and left at -50 °C for 30 min. After that time, the melting was studied by heating the sample at 5 °C/min from -50 °C to 60 °C. At least duplicate determinations were carried out. Characteristic data were obtained using the Shimadzu DSC-60A plus software.

2.2.5. Color

Clarified butter was measured at 50 °C using a Minolta Chroma-Meter CR-400 colorimeter. The parameters L*, a*, and b* were recorded in triplicate (Chiyodaku, Tokyo, Japan).

2.3. Statistical Analysis

For each season, data on fat content, FAP, and nutritional quality indices (AI, H/H, n-6/n-3) in milk were analyzed using repeated measures ANOVA with PROC GLIMMIX of SAS 9.04 (SAS Institute Inc., Cary, NC, USA). The statistical models included treatment (HP and LP), period (1 to 5), and the interaction between treatment and period. Farms within each treatment were considered as random effects. The Kenward-Roger approximation was utilized to calculate the denominator degrees of freedom for the fixed effects tests of the model. Least squares (LS) means were generated using the LSMEANS/DIFF option, and post hoc comparisons were conducted with the Tukey test.
For butter variables (FAP, SI, hardness, and color), analyses were performed using one-way ANOVA, followed by post hoc Tukey tests, to compare the two contrasting farms (GRZ and C) in each season. Results were considered significant at p ≤ 0.05. Data are presented as mean ± SEM (standard error of the mean). The temperature and fusion enthalpy variables obtained from the DSC were analyzed using descriptive statistics. The mean ± SD data for both variables are shown. All variables were performed using SAS software 9.04. (SAS Institute Inc., Cary, NC, USA).

3. Results

3.1 Milk fat and fatty acid profile in commercial dairy farms. In spring, the HP treatment had a milk fat content of 3.71 ± 0.06%, and the LP had 3.78 ± 0.06%. In fall the HP had fat values of 3.84 ± 0.08%, and LP had fat values of 3.89 ± 0.08%. In spring, there were no treatment or period effects, and no interaction between the treatments and period was observed. In fall, there was no treatment effect and no interaction between treatments and period but there was a period effect. In period 1, the milk fat content was 4.08 ± 0.10%, whereas in period 2, it was 3.65 ± 0.10% (p = 0.037).
There were no differences between treatments (HP and LP) in total SFAs or the total sum (Table 4) in both seasons. On the other hand, SFAs had the highest percentage (Table 4) compared to MUFAs and PUFAs (Tables 6 and 7). Regarding SFAs, it was observed that about 10% were short-chain (especially butyric acid 4:0 and caproic acid 6:0), and the three most abundant fatty acids in this lipid fraction of milk were palmitic acid 16:0, myristic acid 14:0, and stearic acid 18:0. Additionally, the presence of branched- chain FAs (BCFAs) was noted, with the most prevalent being the 15:0 and 17:0 (odd- chain FAs; Table 5).
There was a treatment effect on most of the iso and anteiso BCFAs (p < 0.05; Table 5) in both seasons (spring and fall). In fall, the BCFAs were higher in HP compared to LP (p = 0.007). Among the BCFAs, the odd- and branched- chain FAs (OBCFA) 15:0 anteiso, 17:0 iso, and 17:0 anteiso were in higher proportions (Table 5). The HP treatment showed a higher proportion of iso FAs compared to the LP (p < 0.05), particularly in spring, while in fall this was observed only for 15:0 (p = 0.003; Table 5).
There were no significant differences in MUFA content between treatments in either season (Table 6). Oleic acid (18:1 cis) was the most abundant of MUFA, with a content ranging from 19.68% to 21.74% in both HP and LP, across both seasons. For vaccenic acid (18:1 t11, TVA), there was a difference between treatments in spring (p = 0.012). The HP treatment had a higher proportion of TVA (4.11 % vs. 2.36 %) compared to LP (Table 6). Regarding total PUFAs, in spring, there were no differences in linoleic acid (18:2 cis) content between treatments (HP and LP); however, in fall, LP had higher total PUFAs than HP and 18:2 cis tended to be higher than HP (Table 7). The content of linolenic acid (18:3 n-3 cis) was higher in HP compared to LP in spring (p = 0.01). Regarding CLA content, milk from HP was higher than LP in both spring (p = 0.014) and fall (p = 0.033, Table 7).
Regarding the indicators of nutritional quality, the atherogenic index (AI) showed no significant differences between the different treatments (HP vs. LP) in either spring or fall. The values obtained for HP were 3.1 ± 0.2 and 3.3 ± 0.3 in spring and fall, respectively, while LP presented values of 2.9 ± 0.2 and 2.7 ± 0.3 in spring and fall, respectively. For the H/H index, we also found no differences between treatments in either season. The values obtained for HP were 0.5 ± 0.03 in both spring and fall, while LP presented values of 0.5 ± 0.03 and 0.6 ± 0.07 in spring and fall, respectively. Regarding the n-6/n-3 ratio, in fall, the HP was lower than the LP treatment (2.8 ± 0.4 vs. 5.8 ± 0.4; p = 0.05), while in spring, there were no differences between treatments.

3.2. Butter production from two farms (GRZ and C)

3.2.1. Fatty acid profile in Butter

In spring, the butter made from GRZ had higher content of CLA, 18:2 cis, TVA, and lower total MUFA content, primarily represented by 14:1, 16:1 compared to C (Table 8). Regarding SFA, although there were no differences in total content, GRZ butter had a higher proportion of 11:0, 12:0 and branched-chain fatty acids 14:0 iso, 15:0 anteiso compared to C (Table 8). In fall, GRZ butter showed a higher proportion of total SFA and BCFA compared to C, showing differences in almost all individual SFAs (Table 8). Regarding unsaturated fatty acids, GRZ butter had lower MUFA and PUFA compared to C, primarily represented by TVA, 18:1 cis, and 18:2 cis (Table 8). The fatty acids that showed differences between treatments (GRZ vs. C) in butter, both in spring and fall, were consistent with those observed in MilkB.

3.2.2. Techno-Functional Characteristics of Butter from Two Farms (GRZ and C)

  • Butter Hardness
There were no differences in firmness between GRZ and C in spring. However, in fall, butter from C showed lower firmness compared to GRZ butter (p = 0.008; Figure 1).
  • Differential Scanning Calorimetry (DSC)
The thermograms obtained from the Differential Scanning Calorimeter (DSC) are shown in Figure 2, for anhydrous milk fat from GRZ and C. The thermal behaviors are very similar, exhibiting two peaks that can be associated with groups of triglycerides with different melting ranges [60]. These are represented by the two peaks observed in the respective thermograms.
The temperature for the peak corresponding to the triglyceride group with the lower melting point (peak 1) ranged from 14.9 to 16.7 °C, while for the triglycerides with the higher melting point (peak 2), the temperature ranged from 32.7 to 34.1 °C (Table 9). Additionally, the enthalpies of fusion for the anhydrous milk fat ranged between 89.4 J/g and 112.9 J/g, as shown in Table 9.
The percentage solids curves for each of the anhydrous milk fats (Figure 3A) show very little difference between them. However, significant differences were observed at -5 °C and between 9 °C and 20 °C, where the percentage solids decreased rapidly (Figure 3B). It is worth noting that firmness and the percentage solids values at 10 °C, (the temperature at which firmness was determined), showed the same trend. Anhydrous milk fat Fall-C, which exhibited the lowest firmness at 10 °C, also had the lowest percentage of solids at that temperature. Conversely, Fall-GRZ anhydrous milk fat, which showed the highest firmness, had the highest percentage of solids compared to the other samples (Figure 3B), consistent with its higher firmness (Figure 1).
  • Color
Regarding color parameters, no differences were observed between GRZ and C butters in spring. In fall, there were no differences in L and a* parameters. Nevertheless, butters from GRZ had higher b* values than those from C (p = 0.003; Table 10).

4. Discussion

4.1. Fatty Acid Profile in Milk

When it comes to the proportion of the main groups of fatty acids composing milk fat, our results align with findings reported by other authors, where 60-70% corresponds to saturated fatty acids (SFA), 20-25% to monounsaturated fatty acids (MUFA), and 3-4% to polyunsaturated fatty acids (PUFA) [36,61,62]. Among the SFAs, the most abundant found in this study were palmitic acid (16:0), myristic acid (14:0), and stearic acid (18:0), which also coincides with reports from other authors in similar dairy cow production systems [63,64]. The fact that these are the most abundant SFAs in cow’s milk is related to the biosynthesis mechanisms of fat in the dairy cow. Lauric acid (12:0), myristic acid (14:0), and palmitic acid (16:0) are primarily synthesized in the mammary gland from precursors originating in the rumen [65]. Particularly, palmitic acid (16:0) is one of the most abundant, partly due to its mixed origin, as it can be synthesized in the gland or come from body reserves [66]. On the other hand, long-chain saturated fatty acids such as stearic acid (18:0) reach the milk primarily from the rumen, originating from fatty acids in the diet that undergo biohydrogenation by ruminal microbiota or from body reserves [65].
While no differences were found in the total amount of SFA between treatments, we observed that milk from HP had a higher proportion of branched- chain FAs (BCFA) compared to LP treatment. Within BCFA, odd- and branched- chain FAs (OBCFA) such as 15:0 and 17:0 iso and anteiso were the most abundant, consistent with findings reported by Ran Ressler et al. [67] for bovine milk. The fact that HP had a higher proportion of these fatty acids compared to LP treatment (both in spring and fall) could be explained by the higher forage to concentrate ratio (F:C ratio) in the diet of HP. This higher F:C ratio could increase the concentration of cellulolytic bacteria in the rumen, which are associated with higher proportions of OBCFA in milk (mainly iso forms), as reported by Vlaeminch et al. [20]. Xin et al. [68] also reported that cellulolytic bacteria had a stronger correlation with OBCFA compared to amylolytic microbiota. Specifically, these authors found a positive correlation between the population of Ruminococcus flavefaciens and the concentrations of 15:0. Based on the reported benefits of OBCFA (specifically 15:0 and 17:0) for human health and the limited knowledge available to date on their levels in commercial dairy systems, this study provides relevant information on the positive association between higher pasture inclusion levels in the diet and OBCFA concentrations in milk, consistent with the authors cited above. This reinforces the importance of including pasture in cows’ diets and the resulting benefits on the quality of milk fat.
Furthermore, the higher proportion of TVA, 18:3 n-3 cis, and CLA (main rumenic FA: 18:2 cis 9 trans 11) in milk from the HP treatment compared to LP, particularly in spring, is consistent with findings reported by Elgersma, Alothman, and Moscovici Joubran [36,62,69], and could be associated with the higher proportion of pasture (percentage of total diet) in HP compared to LP treatment during this season (Table 2). As widely reported, fresh pastures contain a high proportion of α-linolenic acid (the main precursor of TVA and CLA in milk) [69] and exhibit maximum digestibility between August to October (spring) [70,71,72], which could explain these differences between systems, although no differences were observed in the total MUFA and PUFA. Taking into account that these FAs constitute the main components of dairy fat considered beneficial for consumers due to their anti-atherogenic, anti-carcinogenic, anti-inflammatory, immunostimulant, and insulin resistance-modulating effects [73], and that current nutritional trends promote an increase in PUFA and CLA consumption [65], milk from HP could be considered healthier than milk from LP. Moreover, regarding the content of TVA and CLA in spring, it is observed that the trends in their variation are similar: increases in their content correspond to increases in the other. This confirms the metabolic dependence for the formation of both fatty acids, due to the stages of biohydrogenation in the rumen and dehydrogenation in the mammary gland [74]. Regarding the indices related to the risk of foods causing cardiovascular problems, the HP treatment showed lower n-6/n-3 ratios compared to LP, both in fall and spring, suggesting that farms with higher pasture inclusion in the diet produce healthier milk (values below 4/1) [23]. While this study did not find differences in AI and H/H between treatment or seasons in milk, the values were similar to those reported by other authors [64,75,76,77].
Overall, we could suggest that milk from farms with a higher proportion of pasture may have health benefits compared to milk from farms with a lower proportion of pasture, as indicated by the higher levels of TVA, CLA and 18:3 n-3 cis in milk fat, together with the higher levels of OBCFA mentioned above. Furthermore, the higher content of OBCFA (especially 15:0 and 17:0), which have demonstrated beneficial effects on human health [17,18], contributes to this suggestion. Therefore, quantifying these fatty acids in milk (especially the latter) from commercial farms in contrasting seasons based on dietary structure is crucial for valorizing attributes related to the nutritional quality of the milk produced.

4.2. Fatty Acid Profile in Butter

Based on the fatty acid profiles obtained from the butter, the higher proportion of CLA and TVA in the GRZ during spring indicates a healthier nutritional profile compared to C. This could be attributed to the higher percentage of pasture (grazing) in the total diet of GRZ cows compared to C. Although there was no difference in 18:3 in the butter, it was observed that this fatty acid was higher in MilkB from GRZ compared to C, which aligns with Elgersma [69], who reported that pasture contributes significantly to 18:2 and 18:3, precursors of major n-3, CLA, and TVA in milk and dairy products.
In the case of fall butter, contrary to expectations, both MilkB and butter from GRZ had a higher proportion of SFA (mainly represented by 14:0, which is considered one of the main atherogenic fatty acids). However, the concentration of OBCFA, such as 15:0 and 17:0 iso and anteiso, especially 17:0 anteiso, was more than twice as high in GRZ compared to C. This is interesting because although GRZ butter had high SFA content, a significant portion consists of OBCFA known for its bioactive properties [78]. These compounds could potentially counteract the adverse effects associated with consuming atherogenic fatty acids found in this dairy product.

4.3. Physical Properties of Butter

The lower firmness observed in butter made from C compared to GRZ in fall aligns with the FAP in MilkB and the butter made. The C treatment had higher PUFA than GRZ and showed a tendency to be higher in MilkB. This increase was mainly explained by high values of 18:2 both in MilkB and butter. Additionally, the C treatment had lower SFA than GRZ in MilkB and butter. While a higher concentration of PUFA was expected in the GRZ treatment compared to C (as per the design with and without fresh pasture in the diet), analysis of the FAP of both treatment’ diets and ingredients revealed that the C treatment, despite lacking grazing, had more than double the 18:2 n-6 and 18:1 n-9 in the diet compared to GRZ. These results could be explained by the high proportion of maize grain and soybean expeller (as concentrate) in the C diet during the butter production periods, as these ingredients provide a relatively high proportion of fat [79], specifically a high proportion of 18:2 n-6 in the diet. In this regard, Roy et al. [80] reported that maize silage generally contains between 30% to 40% grain, which is rich in 18:2 n-6, 18:1 n-9, and low in 18:3 n-3. Regarding the addition of soybean in pasture-based systems, it has been reported to decrease FAs from 6:0 to 14:0 and increase 18:0, 18:1, 18:2, and 18:3 in milk [81,82]. Therefore, the C producer‘s diet, composed of maize grain and soybean expeller, had a high contribution of 18:2 in the diet. On one hand, this may have increased the passage rate and resulted in higher concentrations of 18:2 in milk [83] . On the other hand, the high concentrations of these FAs in the rumen may have inhibited ruminal biohydrogenation, resulting in lower levels of SFA and higher levels of MUFA and PUFA in MilkB and butter from the C farm. Furthermore, the higher levels of MUFA and PUFA could explain the lower firmness of C butter compared to those from GRZ [25,84].
The high firmness of butter from GRZ in fall could be explained by the higher proportion of SFA compared to C, considering that short-chain fatty acids (with low melting points) did not vary significantly between samples. This is observed not only in total SFA, but also in most individual SFAs. It has been reported that butter made from creams with a high proportion of SFA are firmer and less spreadable than those made from creams with a higher proportion of UFA [25,85], determined by the higher melting point of SFAs [86]. Among SFAs, palmitic acid (16:0), which has a high melting point (62.9 °C), was higher in GRZ butter than in C butter, leading to a higher spreadability index, consistent with findings reported by Techeira et al. [84], Marangoni and Ghazani[85] and Chamberlain et al. [87], and therefore higher firmness. In spring, no differences in butter firmness were observed between GRZ and C. These results could be explained because MilkB and butters from the two farms (GRZ and C) had a similar FAP in this season, with no differences found in most FAs (mainly SFA and PUFA)
Through DSC analysis, the melting behavior, melting temperatures, enthalpies of fusion, and solids content at different temperatures was investigated. Table 9 shows the temperatures corresponding to the peak for triglycerides with lower melting points (ranging between 14.9 to 16.7 °C) and for those with higher melting points (ranging between 32.7 to 34.1 °C). In this study, differences in the feeding regimes between GRZ and C, in fall, led to different enthalpy of fusion values for the anhydrous milk fats used in the butter made from GRZ versus C. This differed from what had occurred in spring, where changes in SFA, MUFA, and PUFA content were not as significant between treatment. The values obtained ranged between 89.4 J/g (Fall-C) and 112.9 J/g (Fall-GRZ). Tomaszewska-Gras [88] reported enthalpy of fusion values for anhydrous milk fat of 80.3 J/g. Although this value is lower than that found in the present study, it is likely due to differences in its composition. The differences in the enthalpy of fusion values found may be attributed to variations in the triacylglycerol composition among different anhydrous milk fats. Additionally, the enthalpy of fusion of fat materials not only depends on the triacylglycerol composition but also on the minor components present in them [89]. The final melting temperature (Tendset) for anhydrous milk fat ranged from 36.4 °C to 42.4 °C. Additionally, DSC analysis revealed that GRZ anhydrous milk fat in fall had the highest solids content (Figure 3B), which correlates with it being the sample with the greatest firmness (Figure 1). The results indicated that most of the variation in composition and thermal properties occurred in the fall samples, likely due to the feeding regimes, specifically, GRZ versus C. For example, in fall, GRZ had a higher SFA content compared to C (21.6% vs. 18.1%) and a lower MUFA content (14.4% vs. 26.5%). Therefore, variations in feeding should be considered as they can affect the physicochemical properties of the butters produced. Consequently, the results shown in Figure 3 were expected because a higher solids content should be reflected in a fatty material with greater firmness.
Regarding the color of the butters, there were differences between GRZ and C in those made in fall. The higher intensity of the yellow color in the butter made from GRZ compared to C could be attributed to pastures having a higher concentration of β-carotene than ensiled feeds, as the ensiling process negatively impacts the concentration of this compound in the feed provided [90]. Since β-carotene is one of the main components influencing butter color, a higher intake of β-carotene in the diet could have resulted in a greater intensity of yellow color in the GRZ butters, consistent with findings reported by O‘Callaghan et al. [25] .
The predominant use of pastures (fresh pasture: grazing or conserved forage) promotes an increase in unsaturated fatty acids and branched- chain fatty acids of nutritional and technological interest (CLA, TVA in spring, 15:0 iso and anteiso: 17:0 iso) in milk. This not only enhances the added value of milk produced in pasture-based systems but also the products such as butter made from it. In this study, samples from actual producers supplying the industry were analyzed, representing commercial production systems and collectively including variations in herd management, pasture availability, and the nature of supplementation. This wide variability in sources makes analyzing the results a complex task, but it also provides a truthful view of the production conditions present in the western coastal systems of the Uruguay. Observing the results for butters produced on a pilot scale highlights the importance of the availability and management of different supplements, pasture levels and season. In the butters made in fall, the reported 18:2 cis values in the feed from farm C doubled those of GRZ, likely due to the nature of the lipids provided by corn grains and the presence of soybean expeller in the supplement. Consequently, in fall, the butters produced from C had lower firmness than those manufactured from GRZ., paradoxically to the inclusion of pasture. In this case, for C, the richness in lipids from the supplement with soybean expeller combined with corn grain in the TMR promoted functional and technological properties associated with high pasture conditions, although it did not increase the yellow coloration or the content of OBCFAs. The texture and crystallization profile of the fats present in the butter then highlight the importance of selecting the overall feed based on the available sources and the possibility of obtaining suitable technological properties even in the absence of pastures

5. Conclusions

The results obtained in this study, conducted on commercial farms, underscore the benefits of high pasture inclusion (in both spring and fall) in the diet of dairy cows for producing healthier milk for consumers. This was evidenced by higher concentrations of TVA, CLA, and 18:3 n-3, and a lower n-6/n-3 ratio. Additionally, higher levels of some SFA, recently reported beneficial for human health, such as OBCFAs 15:0 and 17:0, were observed. Moreover, butter derived from milk of pasture-based system (GRZ) also exhibited a healthier profile, characterized by higher proportions of CLA and TVA in spring, and higher proportions of 15:0 and 17:0 iso and anteiso in fall butters. In addition, yellow color of GRZ butters was stronger. However, contrary to expectations, GRZ butters in fall presented greater firmness, possibly related to the higher palmitic/oleic ratio, influenced by the fatty acid composition of the diet. This demonstrates the potential benefits of adjusting the management of feed for confined cows with oils and highlights the need for further studies considering supply and cost constraints.

Author Contributions

Conceptualization: E. K., L.G., I. V., A.G., S. J., T.L-P. Methodology: M.R., S.J., E.K., E.B., A.G., I.V., B.I., MN.M., L.G., T.L-P. Software: A.G., B.I., S.J. Validation: L.G., I.V., T.L-P. formal analysis: A.G., L.G., I.V., T.L-P. investigation I.V., S.J., E.K., A.G., L.G., T.L-P., M N.M., E.B., M.R., B.I. resources: L.G., T.L-P. data curation: A.G. writing—original draft preparation, L.G., I.V., T.L-P writing—review and editing: L.G., I.V., T. L-P, MN.M, A.G. visualization, L.G., B.I., I.V., A.G. supervision, T. L-P, I.V. project administration, L.G., T. L-P. funding acquisition: L.G., T. L-P. All authors have read and agreed to the published version of the manuscript.

Funding

This research was funded by Unidad Tecnológica de Lácteos, Departamento de Alimentos, Universidad Tecnológica del Uruguay (UTEC), Grupo de Investigación Estratégica (GIE)- UTEC, Comisión Sectorial de Investigación Científica- Universidad de la República (CSIC-Udelar).

Institutional Review Board Statement

Not applicable.

Informed Consent Statement

Not applicable.

Data Availability Statement

The original contributions presented in the study are included in the article material, further inquiries can be directed to the corresponding author/s.

Acknowledgments

This work would not have been possible without the contributions of farmers associations from the West Coast of Uruguay, such as “Mesa Lechera del Litoral” and the “Asociación de Productores de Leche de Paysandú” as well as the undergraduate students. The authors would also like to thank Priscila Guinovart for her English proofreading contribution.

Conflicts of Interest

The authors declare no conflicts of interest.

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Figure 1. Butter hardness elaborated from GRZ: pasture- based system (grazing + supplement); C: confined (total mixed ration, TMR). Difference between farms (GRZ and C) in spring and fall are shown with small letter and capital letter respectively (p < 0.05).
Figure 1. Butter hardness elaborated from GRZ: pasture- based system (grazing + supplement); C: confined (total mixed ration, TMR). Difference between farms (GRZ and C) in spring and fall are shown with small letter and capital letter respectively (p < 0.05).
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Figure 2. Thermograms of anhydrous milk fat from the prepared butters from GRZ: pasture-based system (grazing + supplement); C: confined (total mixed ration, TMR), in spring and fall.
Figure 2. Thermograms of anhydrous milk fat from the prepared butters from GRZ: pasture-based system (grazing + supplement); C: confined (total mixed ration, TMR), in spring and fall.
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Figure 3. (A) Percentage of solids in anhydrous milk fat from butters produced under GRZ: pasture-based system (grazing + supplement) and C: confined (total mixed ration, TMR) in spring and fall. (B): Percentage of solids in anhydrous milk fat: magnification in the range between 8 and 13 °C, with an arrow indicating the temperature at which firmness was determined (10 °C).
Figure 3. (A) Percentage of solids in anhydrous milk fat from butters produced under GRZ: pasture-based system (grazing + supplement) and C: confined (total mixed ration, TMR) in spring and fall. (B): Percentage of solids in anhydrous milk fat: magnification in the range between 8 and 13 °C, with an arrow indicating the temperature at which firmness was determined (10 °C).
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Table 1. Average values of neutral detergent fiber (NDF), acid detergent fiber (ADF), and ether extract (EE) for each group (pasture and supplement) in Spring and Fall.
Table 1. Average values of neutral detergent fiber (NDF), acid detergent fiber (ADF), and ether extract (EE) for each group (pasture and supplement) in Spring and Fall.
Spring Fall
HP LP HP LP
Past Suppl Total diet** Past* Suppl Total diet** Past Suppl Total diet** Past* Suppl Total diet**
NDF 47.3±
4.8		 30.3±
7.3		 43.1±
3.5		 46.3±
1.7		 36.2±
1.8		 37.2±
2.6		 36.7±
6.9		 43.9±
7.6		 40.1±
5.7		 36.9±
2.4		 44.7±
5.8		 44.5±
5.9
ADF 26.2±
2.3		 14.3±
6.1		 23.4±
1.5		 26.3±
1.5		 19.8±
2.5		 20.6±
2.2		 20.1±
5.0		 24.1±
6.5		 22.0±
3.6		 20.9±
1.7		 26.5±
7.2		 26.5±
7.2
Lipids 3.7±
0.9		 4.8±
1.6		 4.1±
1.1		 2.6±
0.06		 3.9±
0.3		 4.2±
0.5		 6.6±
4.1		 3.9±
1.2		 5.4±
2.5		 6.9±
0.1		 5.8±
1.5		 5.8±
1.5
Past: Pasture; Supp: Supplement. HP: High Pasture; LP: Low Pasture. NDF: Neutral Detergent Fiber. ADF: Acid Detergent Fiber. * The “LP pasture” data in both seasons corresponds to the only producer of that treatment offering fresh pasture in the diet. ** The total diet chemical composition value was calculated by weighting the inclusion level of pasture and supplement.
Table 2. Fatty acid profiles of pasture and supplement in both treatments: HP and LP in Spring and Fall.
Table 2. Fatty acid profiles of pasture and supplement in both treatments: HP and LP in Spring and Fall.
Spring Fall
Fatty acids (g/100 g fat) HP LP HP LP
Past Suppl Past Suppl Past Suppl Past Suppl
10:0 0.36 ± 0.20 0.11 ± 0.03 0.82 ± 0.44 0.41 ± 0.22 0.06 ± 0.05 0.03 ± 0.05 0.20 ± 0 0.29 ± 0.18
12:0 0.74 ± 0.21 0.14 ± 0.05 1.3 ± 0.55 0.61 ± 0.27 0.36 ± 0.08 0.34 ± 0.27 0.30 ± 0 0.45 ± 0.42
14:0 2.09 ± 0.79 0.50 ± 0.15 4.62 ± 1.53 1.69 ± 0.70 0.95 ± 0.31 0.56 ± 0.23 0.63 ± 0.05 1.29 ± 1.08
16:0 18.25 ± 2.91 15.23 ± 2.19 24.82 ± 2.36 16.49 ± 1.73 12.6 ± 1.5 15.3 ± 1.4 12.4 ± 0.1 14 ± 3.1
16:1 cis 0.37 ± 0.19 0.17 ± 0.06 0.60 ± 0.27 0.33 ± 0.08 0.03 ± 0.08 0.29 ± 0.21 0.18 ± 0.05 0.29 ± 0.37
17:0 0.36 ± 0.27 0.11 ± 0.03 0.44 ± 0.05 0.13 ± 0.12 0 ± 0 0 ± 0 0.05 ± 0.10 0.03 ± 0.05
18:0 3.1 ± 0.97 2.06 ± 0.46 5.24 ± 0.33 3.49 ± 0.42 2.21 ± 0.32 2.49 ± 0.48 2.08 ± 0.05 4.35 ± 1.60
18:1 trans-vaccenic 0.25 ± 0.13 0.05 ± 0.08 0.52 ± 0.16 0.12 ± 0.13 0.11 ± 0.07 0.13 ± 0.13 0.35 ± 0.10 0.08 ± 0.09
18:1 cis 7.21 ± 4.19 28.81 ± 6.01 11.44 ± 1.15 22.8 ± 2.76 5.00 ± 1.57 27.68 ± 5.71 3.90 ± 0.60 22.66 ± 2.50
18:2 trans 1.03 ± 1.00 0 ± 0 0.58 ± 0.11 0.07 ± 0.10 0.20 ± 0.19 0.13 ± 0.10 0.15 ± 0.10 0.22 ± 0.20
18:2 cis 13.41 ± 5.05 46.43 ± 4.86 10.18 ± 0.38 41.37 ± 4.35 13.77 ±3.04 41.81 ± 5.82 14.18 ± 1.35 37.90 ± 10.02
20:0 0.79 ± 0.39 0.43 ± 0.05 0.58 ± 0.11 0.49 ± 0.14 0.38 ± 0.08 0.62 ± 0.14 0.40 ± 0 0.53 ± 0.21
20:1 0.04 ± 0.08 0.39 ± 0.14 0 ± 0 0.11 ± 0.09 0.10 ± 0.10 0.49 ± 0.09 0 ± 0 0.19 ± 0.18
18:3 cis 42.42 ± 12.93 3.41 ± 1.66 31.72 ± 11.61 8.36 ± 4.11 58.66 ± 7.61 6.84 ± 5.47 57.60 ± 2.40 13.36 ± 9.32
22:0 0.51 ± 0.12 0.24 ± 0.06 0.54 ± 0.05 0.33 ± 0.14 0.66 ± 0.24 0.53 ± 0.13 0.55 ± 0.10 0.48 ± 0.16
24:0 0.47 ± 0.12 0.3 ± 0.08 0.32 ± 0.16 0.27 ± 0.12 0.40 ± 0.07 0.55 ± 0.25 0.53 ± 0.05 0.38 ± 0.11
SFA 27.21 ± 5.52 19.17 ± 2.64 40.22 ± 7.28 24.73 ± 3.65 18.77 ± 3.60 18.36 ± 1.57 19.25 ± 2.10 18.76 ± 2.01
MUFAs cis 7.63 ± 4.31 29.37 ± 5.89 12.4 ± 1.37 23.25 ± 2.76 19.56 ± 8.65 22.14 ± 15.97 6.68 ± 7.15 8.47 ± 6.18
PUFAs cis 55.83 ± 9.16 49.84 ± 5.56 41.9 ± 11.23 49.73 ± 3.95 56.80 ± 8.91 56.47 ± 16.42 66.80 ± 6.80 67.38 ± 6.93
Trans 1.28 ± 1.08 0.05 ± 0.08 1.1 ± 0.27 0.19 ± 0.19 0.26 ± 0.30 0.15 ± 0.18 0.38 ± 0.25 0.38 ± 0.22
Total fat 3.73 ± 1.00 4.86 ± 1.63 2.62 ± 0.06 3.95 ± 0.30 5.96 ± 1.17 9.03 ± 5.52 5.78 ± 2.00 4.41 ± 0.68
Past: Pasture; Suppl: Supplement. HP: High Pasture; LP: Low Pasture. SFA: Saturated Fatty Acids; MUFAs: Monounsaturated Fatty Acids; PUFAs: Polyunsaturated Fatty Acids.
Table 3. Fatty acid profiles of the total diet (pasture and supplement) from two selected dairy farms for butter production (C and GRZ).
Table 3. Fatty acid profiles of the total diet (pasture and supplement) from two selected dairy farms for butter production (C and GRZ).
Spring Fall
Fatty acids
(g/100 g fat)		 GRZ C GRZ C
16:0 18.5 ± 2.5 17.0 ± 2.1 15.4 ± 1.4 12.6 ± 1.6
18:0 3.7 ± 0.7 3.5 ± 0.3 2.6 ± 0.4 4.0 ± 0.6
18:1 n9 cis 11.3 ± 5.1 18.2 ± 1.9 14.0 ± 3.3 26.2 ± 1.5
18:2 n6 cis 21.7 ± 4.9 34.6 ± 2.3 25.5 ± 4.4 47 ± 5.2
18:3 n3 cis 29.2 ± 7.2 15.1 ± 2.8 32.2 ± 6.5 4.2 ± 5.8
GRZ: pasture-based system (grazing + supplement); C: confined (total mixed ration, TMR).
Table 4. Saturated Fatty Acid Profile of Milk in Spring and Fall.
Table 4. Saturated Fatty Acid Profile of Milk in Spring and Fall.
Fatty Acid (g/100g fat) Spring Fall
T P SEM p-value T P SEM p-value
1 2 3 4 5 T P T*P 1 2 3 4 5 T P T*P
4:0 HP 3.03 2.92 3.54 3.43 2.51 2.76 0.09 ns 0.022 ns HP 3.16 3.20 3.31 2.94 3.49 2.87 0.13 ns ns ns
LP 2.86 2.58 3.18 2.61 2.55 3.37 LP 3.30 3.81 3.13 3.26 3.28 3.04
6:0 HP 1.82 1.93a 2.12a 1.91a 1.49a 1.63b 0.05 ns 0.050 0.045 HP 2.00 1.86 2.15 1.92 2.25 1.82 0.10 ns ns ns
LP 1.86 1.73a 2.04a 1.64a 1.70a 2.21a LP 2.13 2.48 2.04 2.13 2.09 1.90
8:0 HP 1.06 1.18 1.23 1.01 0.87 1.00 0.05 ns ns ns HP 1.23 1.07 1.35 1.24 1.40 1.09 0.08 ns ns ns
LP 1.13 1.10 1.19 1.00 1.04 1.34 LP 1.30 1.47 1.30 1.28 1.31 1.12
10:0 HP 2.37 2.77 2.78 2.15 2.04 2.12 0.16 ns ns ns HP 2.78 2.75 2.93 2.73 3.10 2.39 0.29 ns 0.015 ns
LP 2.64 2.64 2.71 2.41 2.52 2.89 LP 2.62 2.65 2.54 2.69 2.83 2.39
11:0 HP 0.30 0.35 0.34 0.26 0.26 0.28 0.02 ns ns ns HP 0.36 0.31 0.42 0.36 0.41 0.29 0.03 ns ns ns
LP 0.32 0.30 0.33 0.29 0.32 0.39 LP 0.31 0.27 0.35 0.33 0.32 0.27
12:0 HP 2.78 3.26 3.20 2.47 2.50 2.49 0.10 ns ns ns HP 3.22 2.93 3.49 3.23 3.63 2.81 0.42 ns ns ns
LP 3.08 3.07 3.19 2.91 2.89 3.32 LP 2.95 2.91 2.82 3.03 3.23 2.75
14:0 HP 10.61 11.56 11.50 10.08 10.20 9.72 0.42 ns ns ns HP 10.98 11.01 11.16 11.03 11.50 10.21 0.85 ns ns ns
LP 10.86 10.84 10.88 10.75 10.66 11.17 LP 10.15 10.61 9.89 10.21 10.54 9.51
15:0 HP 1.37 1.42 1.36 1.35 1.34 1.38 0.14 ns ns ns HP 1.20 1.17 1.26 1.21 1.23 1.11 0.06 ns ns ns
LP 1.21 1.23 1.19 1.19 1.13 1.29 LP 1.04 1.03 1.04 1.08 1.01 1.02
16:0 HP 29.46 29.75 29.69 29.14 30.00 28.71 1.00 ns ns ns HP 30.75 31.40 30.59 30.43 31.07 30.26 1.16 ns ns ns
LP 29.94 29.30 29.65 30.29 29.73 30.71 LP 28.79 30.00 28.41 28.06 29.19 28.28
17:0 HP 0.82 0.79 0.81 0.82 0.83 0.87 0.06 ns ns ns HP 0.71 0.64 0.68 0.76 0.73 0.76 0.04 ns ns ns
LP 0.69 0.71 0.69 0.69 0.65 0.71 LP 0.67 0.62 0.70 0.68 0.64 0.72
18:0 HP 10.08 9.27 9.73 10.72 10.24 10.42 0.45 ns ns ns HP 9.15 9.12 8.52 9.59 8.56 9.95 0.85 ns ns ns
LP 10.10 11.03 10.49 9.92 10.12 8.92 LP 10.93 11.19 11.44 10.57 10.37 11.09
Total SFA HP 63.88 65.36 66.46 63.55 62.45 61.59 1.08 ns ns ns HP 65.55 64.94 66.02 65.59 67.51 63.71 2.13 ns ns ns
LP 64.91 64.71 66.01 63.87 63.49 66.46 LP 64.34 67.19 63.84 63.46 64.95 62.24
HP= High pasture; LP= Low pasture. T: treatments; P: periods, T*P: interaction between treatments and periods. SFA: saturated fatty acid. SEM: standard error of the mean. Different letters indicate significant difference (p < 0.05). ns: no significance.
Table 5. Branched- chain Fatty Acid Profile of Milk in Spring and Fall.
Table 5. Branched- chain Fatty Acid Profile of Milk in Spring and Fall.
Fatty Acid (g/100g fat) Spring Fall
T P SEM p-value T P SEM p-value
1 2 3 4 5 T P T*P 1 2 3 4 5 T P T*P
14:0 iso HP 0.17 0.16 0.15 0.18 0.17 0.19 0.01 ns ns ns HP 0.13 0.13 0.12 0.14 0.12 0.12 0.01 ns ns ns
LP 0.13 0.14 0.12 0.13 0.11 0.13 LP 0.11 0.10 0.12 0.11 0.11 0.10
15:0 iso HP 0.36a 0.36 0.35 0.36 0.33 0.37 0.02 0.029 ns ns HP 0.30a 0.27 0.31 0.32 0.29 0.30 0.01 0.003 ns ns
LP 0.28b 0.30 0.32 0.25 0.24 0.27 LP 0.24b 0.25 0.24 0.23 0.22 0.24
15:0 anteiso HP 0.69a 0.74 0.71 0.70 0.64 0.68 0.03 0.008 ns ns HP 0.56a 0.52 0.57 0.58 0.56 0.56 0.01 0.0004 ns ns
LP 0.48b 0.49 0.48 0.48 0.46 0.51 LP 0.43b 0.45 0.43 0.44 0.43 0.42
16:0 iso HP 0.34a 0.35 0.34 0.34 0.33 0.35 0.01 0.003 ns ns HP 0.29 0.26 0.27 0.31 0.30 0.30 0.02 ns ns ns
LP 0.30b 0.33 0.29 0.30 0.29 0.31 LP 0.28 0.25 0.30 0.30 0.28 0.28
17:0 iso HP 0.43a 0.37 0.46 0.45 0.44 0.44 0.02 0.005 ns ns HP 0.38 0.34 0.38 0.40 0.37 0.44 0.02 ns 0.003 ns
LP 0.34b 0.36 0.35 0.34 0.33 0.33 LP 0.33 0.29 0.35 0.34 0.30 0.37
17:0 anteiso HP 0.53 0.55 0.53 0.52 0.50 0.55 0.03 ns ns ns HP 0.48 0.43 0.48 0.50 0.50 0.48 0.02 ns ns ns
LP 0.44 0.46 0.43 0.44 0.43 0.47 LP 0.43 0.38 0.46 0.45 0.41 0.46
18:0 iso HP 0.054a 0.057 0.053 0.052 0.052 0.057 0.001 0.025 0.040 ns HP 0.063 0.059 0.053 0.060 0.067 0.077 0.01 ns ns ns
LP 0.048b 0.051 0.048 0.046 0.046 0.050 LP 0.045 0.045 0.048 0.043 0.039 0.050
Total BCFA HP 2.57a 2.58 2.59 2.59 2.46 2.64 0.09 0.006 ns ns HP 2.19ª 2.01 2.18 2.30 2.21 2.27 0.06 0.007 0.030 ns
LP 2.02b 2.13 2.03 1.98 1.91 2.06 LP 1.86b 1.77 1.95 1.92 1.78 1.91
HP= High pasture; LP= Low pasture. T: treatments; P: periods, T*P: interaction between treatments and periods. BCFA: branched- chain fatty acid. SEM: standard error of the mean. Different letters indicate significant difference (p < 0.05). ns: no significance.
Table 6. Monounsaturated Fatty Acid Profile of Milk in Spring and Fall.
Table 6. Monounsaturated Fatty Acid Profile of Milk in Spring and Fall.
Fatty Acid (g/100g fat) Spring Fall
T P SEM p-value T P SEM p-value
1 2 3 4 5 T P T*P 1 2 3 4 5 T P T*P
14:1 cis HP 0.85 0.99 0.92 0.79 0.81 0.76 0.05 ns ns ns HP 1.01 1.00 1.17 0.99 1.04 0.88 0.11 ns ns ns
LP 0.92 0.89 0.88 0.92 0.84 1.05 LP 0.84 0.76 0.91 0.86 0.87 0.79
16:1 cis HP 1.29 1.34 1.27 1.28 1.35 1.23 0.12 ns ns ns HP 1.59a 1.58 1.71 1.53 1.59 1.54 0.05 0.022 ns ns
LP 1.36 1.39 1.31 1.41 1.23 1.44 LP 1.31b 1.17 1.42 1.33 1.26 1.39
18:1 elaidic HP 0.23 0.26 0.17 0.22 0.26 0.26 0.04 ns ns ns HP 0.22b 0.31 0.22 0.24 0.20 0.11 0.04 0.015 ns ns
LP 0.34 0.32 0.34 0.39 0.41 0.25 LP 0.38a 0.33 0.34 0.41 0.36 0.46
18:1 trans-vaccenic HP 4.11a 4.14 3.84 4.22 4.24 4.10 0.28 0.012 ns ns HP 3.55 3.80 3.45 3.44 3.39 3.70 0.41 ns ns ns
LP 2.36b 2.28 2.35 2.58 2.38 2.20 LP 2.40 1.91 2.41 2.58 2.34 2.73
18:1 cis HP 19.68 18.00 18.04 19.97 21.69 20.68 0.73 ns ns ns HP 19.68 20.68 18.65 19.43 18.01 21.62 1.83 ns ns ns
LP 21.04 21.32 20.54 21.81 22.23 19.29 LP 21.75 20.77 22.78 21.27 21.01 22.94
Total MUFA cis HP 21.82 20.34 20.23 22.04 23.84 22.67 0.68 ns ns ns HP 22.28 23.26 21.52 21.94 20.65 24.04 0.73 ns ns ns
LP 23.32 23.61 22.73 24.14 24.30 21.79 LP 23.90 22.70 25.11 23.45 23.13 25.12
HP= High pasture, LP= Low pasture. T: treatments, P: periods, T*P: interaction between treatments and periods. MUFA: monounsaturated fatty acid. SEM: standard error of the mean. Different letters indicate significant difference (p < 0.05). ns: no significance.
Table 7. Polyunsaturated Fatty Acid Profile of Milk in Spring and Fall.
Table 7. Polyunsaturated Fatty Acid Profile of Milk in Spring and Fall.
Fatty Acid (g/100g fat) Spring Fall
T P SEM p-value T P SEM p-value
1 2 3 4 5 T P T*P 1 2 3 4 5 T P T*P
18:2 trans HP 1.12a 1.07 1.05 1.09 1.19 1.19 0.04 0.003 ns ns HP 1.27a 1.22 1.21 1.22 1.24 1.45 0.04 0.031 ns ns
LP 0.89b 0.79 0.79 1.17 0.86 0.82 LP 1.12b 1.00 1.13 1.09 1.04 1.33
18:2 cis HP 1.72 1.44 1.54 1.91 1.80 1.91 0.32 ns ns ns HP 1.48 1.33 1.51 1.55 1.33 1.69 0.08 <.0001 ns ns
LP 2.54 2.42 2.53 2.55 2.64 2.58 LP 2.86 2.45 2.62 2.99 3.07 3.19
18:2 CLA HP 1.65a 1.86 1.51 1.58 1.70 1.62 0.11 0.014 ns ns HP 1.43a 1.53 1.53 1.47 1.37 1.29 0.15 0.033 ns ns
LP 0.89b 0.79 0.83 0.96 0.93 0.94 LP 0.86b 0.66 0.97 0.90 0.85 0.90
18:3 n3 cis HP 0.89a 0.82 0.89 0.97 0.87 0.90 0.08 0.033 ns ns HP 0.53 0.47 0.52 0.56 0.55 0.57 0.03 ns 0.03 ns
LP 0.54b 0.36 0.50 0.63 0.60 0.63 LP 0.50 0.37 0.54 0.54 0.54 0.53
Total PUFA cis HP 2.61 2.27 2.43 2.88 2.67 2.82 0.24 ns ns ns HP 2.02b 1.80 2.03 2.11 1.87 2.26 0.08 <.0001 ns ns
LP 3.09 2.78 3.03 3.17 3.24 3.21 LP 3.37a 2.82 3.16 3.53 3.61 3.72
HP= High pasture, LP= Low pasture. T: treatments, P: periods, T*P: interaction between treatments and periods. PUFA: polyunsaturated fatty acid. SEM: standard error of the mean. Different letters indicate significant difference (p < 0.05). ns: no significance.
Table 8. Fatty Acid Profile of Butter and MilkB Elaborated in Spring and Fall.
Table 8. Fatty Acid Profile of Butter and MilkB Elaborated in Spring and Fall.
Spring Fall
Fatty Acid (g/100g fat) Butter MilkB Butter MilkB
GRZ C SEM p-value GRZ C SEM p-value GRZ C SEM p-value GRZ C SEM p-value
4:0 3.84 2.61 0.58 ns 2.36 2.97 0.44 ns 2.75a 2.62b 0.02 0.043 2.74 2.78 0.22 ns
6:0 2.33 1.70 0.25 ns 1.59 1.82 0.27 ns 1.86a 1.59 b 0.01 0.002 2.00 1.56 0.13 ns
8:0 1.28 1.87 0.57 ns 1.04 1.03 0.13 ns 1.12a 0.92 b 0.01 0.003 1.30 0.91 0.12 ns
10:0 2.8 2.78 0.25 ns 2.52 2.33 0.11 ns 2.71a 1.99 b 0.04 0.005 3.13 1.86 0.32 ns
11:0 0.34a 0.32b 0.002 0.040 0.33 0.34 0.01 ns 0.29a 0.23 b 0.01 0.040 0.38 0.22 0.05 ns
12:0 3.20a 3.00b 0.01 0.006 3.13 2.65 0.25 ns 3.27a 2.33 b 0.02 0.001 3.73 2.16 0.41 ns
14:0 iso 0.18a 0.16b 0.002 0.046 0.14 0.15 0.02 ns 0.11a 0.09 b 0.002 0.020 0.12 0.10 0.01 ns
14:0 11.37 11.06 0.17 ns 11.56 10.14 0.43 ns 11.36a 8.86 b 0.05 0.001 11.83 7.92 0.73 ns
15:0 iso 0.36 0.40 0.01 ns 0.32 0.31 0.03 ns 0.25a 0.19b 0.004 0.009 0.28 0.23 0.01 ns
15:0 anteiso 0.76a 0.67b 0.01 0.012 0.66 0.59 0.04 ns 0.48a 0.39b 0.01 0.010 0.57a 0.43b 0.01 0.004
14:1 0.93b 1.03a 0.01 0.027 0.96 0.95 0.14 ns 0.83 0.72 0.03 ns 0.95 0.66 0.06 ns
15:0 1.55b 1.62a 0.01 0.025 1.50 1.60 0.09 ns 1.16a 0.99b 0.004 0.001 1.34 1.07 0.05 ns
16:0 iso 0.38 0.40 0.01 ns 0.32 0.32 0.03 ns 0.27 0.27 0.003 ns 0.30 0.27 0.01 ns
16:0 29.79 31.14 0.45 ns 33.25 30.99 1.94 ns 34.23a 28.29b 0.43 0.010 32.61a 25.81b 0.58 0.014
17:0 iso 0.48 0.5 0.02 ns 0.43 0.36 0.03 ns 0.45a 0.33b 0.01 0.008 0.38 0.42 0.04 ns
17:0 anteiso 0.63 0.69 0.01 ns 0.53 0.56 0.05 ns 0.42a 0.19b 0.01 0.006 0.48 0.51 0.03 ns
16:1 1.34b 1.71a 0.03 0.016 1.49 1.57 0.17 ns 1.47 1.35 0.02 ns 1.50 1.36 0.10 ns
17:0 0.94b 1.00a 0.01 0.030 0.88 0.86 0.04 ns 0.75a 0.69b 0.01 0.022 0.80 0.82 0.05 ns
18:0 iso 0.1 0.08 0.03 ns 0.06 0.05 0.003 ns 0.06 0.06 0.01 ns 0.06 0.06 0.01 ns
18:0 9.06 8.97 0.26 ns 8.30 9.06 1.27 ns 9.36b 11.16a 0.10 0.006 8.65b 11.81a 0.51 0.048
18:1 elaidic 0.22 0.19 0.03 ns 0.20 0.29 0.04 ns 0.22b 0.41a 0.03 0.035 0.17b 0.50a 0.04 0.035
18:1 trans-vaccenic 3.22a 1.89b 0.15 0.024 3.65 2.33 0.11 0.015 2.89b 2.97a 0.01 0.009 3.14 2.57 0.26 ns
18:1 cis 17.65b 19.95a 0.35 0.043 17.33 20.80 1.85 ns 18.10b 25.73a 0.22 0.002 17.72 27.28 1.83 ns
18:2 trans 0.81 0.90 0.08 ns 1.10 0.88 0.16 ns 1.01b 1.86a 0.05 0.008 1.19 1.34 0.08 ns
18:2 cis 2.00a 1.77b 0.02 0.010 1.90 1.53 0.15 ns 1.70b 2.84a 0.06 0.005 1.74b 3.23a 0.24 0.048
18:2 CLA 1.44a 0.98b 0.07 0.042 1.57 1.21 0.12 ns 0.92 0.91 0.005 ns 1.06 0.97 0.08 ns
18:3 cis 1.09 1.02 0.07 ns 1.07a 0.96b 0.02 0.050 0.56 0.55 0.02 ns 0.60 0.58 0.02 ns
20:0 0.18 0.18 0.003 ns 0.18 0.17 0.03 ns 0.15a 0.14b 0.001 0.011 0.15 0.17 0.01 ns
Total SFA 66.67 66.25 1.40 ns 66.65 63.95 2.01 ns 69.02a 59.79b 0.45 0.005 68.66a 57.09b 1.68 0.040
Total MUFAcis 19.92b 22.68a 0.39 0.037 19.78 23.31 1.54 ns 20.76b 28.17a 0.22 0.002 20.17 29.30 1.86 ns
Total PUFAcis 3.1 2.78 0.08 ns 2.97 2.50 0.15 ns 2.27b 3.38a 0.07 0.008 2.34 3.81 0.26 ns
Total BCFA 2.87 2.90 0.03 ns 2.46 2.34 0.18 ns 2.04a 1.51b 0.02 0.002 2.19 2.01 0.08 ns
Total Trans 4.26a 2.98b 0.20 0.046 4.95 3.51 0.29 ns 4.12b 5.24a 0.08 0.009 4.26 5.20 0.21 ns
Spreadability index (C16:0/C18:1) 1.69 1.56 0.05 ns 1.93 1.52 0.25 ns 1.89b 1.10a 0.04 0.005 1.85 0.95 0.13 0.037
GRZ: pasture- based system (grazing + supplement); C: confined (total mixed ration, TMR). Milk B: milk to butter. SFA: saturated fatty acid. BCFA: branched- chain fatty acid. MUFAcis: monounsaturated fatty acid. PUFAcis: polyunsaturated fatty acid. SEM: standard error of the mean. Different letters indicate significant difference (p < 0.05). ns: no significance.
Table 9. Peak temperature (Tpeak), enthalpies of fusion (ΔH) and final melting temperature (Tendset) for anhydrous milk fats from butters elaborated in Spring and Fall.
Table 9. Peak temperature (Tpeak), enthalpies of fusion (ΔH) and final melting temperature (Tendset) for anhydrous milk fats from butters elaborated in Spring and Fall.
Tpeak (°C) ΔH (J/g) Tendset (°C)
1 2
Spring-GRZ 15.7 ± 0.4 34.1 ± 0.9 96.9 ± 2.9 39.9 ± 0.3
Spring-C 18.2 ± 0.5 33.1 ± 0.8 97.9 ± 3.0 36.4 ± 0.4
Fall-GRZ 17.2 ± 0.8 33.7 ± 0.9 112.9 ± 3.3 37.1 ± 0.3
Fall-C 16.4 ± 0.7 35.9 ± 0.9 89.4 ± 2.6 42.4 ± 0.5
GRZ: pasture-based system (grazing + supplement) and C: confined (total mixed ration, TMR) in spring and fall.
Table 10. Evaluation of color from butter elaborated in Spring and Fall.
Table 10. Evaluation of color from butter elaborated in Spring and Fall.
Spring Fall
GRZ C SEM p-value GRZ C SEM p-value
L* 55.08 57.16 0.58 ns 59.24 58.95 0.14 ns
a* -5.05 -5.09 0.04 ns -4.57 -4.57 0.13 ns
b* 22.88 23.26 1.00 ns 28.39a 15.71b 0.46 0.003
GRZ: pasture-based system (grazing + supplement); C: confined (total mixed ration, TMR). SEM: standard error of the mean. Different letters indicate significant difference (p < 0.05). ns: no significance.
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