Research Article

Multi Feed Supplement Prevents the Impact of Heat Stress on Dairy Cattle at Low Altitudes in Indonesia

Ujang Hidayat Tanuwiria1, Bambang Kholiq Mutaqin1, Amriana Hifizah2, Andi Mushawwir1

1Department of Animal Nutrition and Feed Technology, Faculty of Animal Science, Padjadjaran University, Jl. Ir. Soekarno KM. 21 Bandung-Sumendang, 45363, Jawa Barat, Indonesia; 2Department of Animal Science, UIN Alauddin Makassar. Jl. H.M. Yasin Limpo No. 36, Gowa 92118, South Sulawesi, Indonesia.

Received | June 03, 2026; Accepted | July 15, 2026; Published | July 28, 2026

*Correspondence | Ujang Hidayat Tanuwiria, Department of Animal Nutrition and Feed Technology, Faculty of Anim. Sci, Padjadjaran University, Jatinangor-Sumedang, West Java 45363, Indonesia; Email: [email protected]

Citation | Tanuwiria UH, Mutaqin BK, Hifizah A, Mushawwir A (2026). Multi feed supplement prevents the impact of heat stress on dairy cattle at low altitudes in Indonesia. Adv. Anim. Vet. Sci., 14(8):1605-1613.

DOI | https://dx.doi.org/10.17582/journal.aavs/2026/14.8.1605.1613

ISSN (Online) | 2307-8316

Copyright: 2026 by the authors. Licensee ResearchersLinks Ltd, England, UK.

This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).

INTRODUCTION

Friesian Holstein (FH) dairy cows are homeothermic animals that require an optimal ambient temperature to live comfortably and produce milk (Ahmed-Farid et al., 2021). The comfort zone for European dairy cows ranges from 13-18℃ (Razzaghi et al., 2022). Meanwhile, Indonesia, especially in lowland areas, has an environmental temperature range of 28-35 ℃ and air humidity of 85% (Tanuwiria et al., 2022). These climatic differences in husbandry can affect livestock’s physiological condition and productivity, thus requiring monitoring to ensure their survival.

Maintaining dairy cows above their comfort zone can cause heat stress, which damages their physiology and blood hematology (Tanuwiria et al., 2022) and alters total leukocyte counts, leukocyte differential counts, and immune and inflammatory responses (Tanuwiria et al., 2025). Leukocyte levels, including neutrophils, lymphocytes, monocytes, eosinophils, and basophils in the blood, serve as indicators for evaluating heat stress experienced by dairy cows. Additionally, heat stress can disrupt physiological and metabolic functions, leading to excessive organ activity, increased inflammation (Wang et al., 2022), and cell damage and death. This affects blood metabolites such as alkaline phosphatase, creatinine, creatine kinase, gamma-Glutamyl Transpeptidase (γ-GT), and Lactate dehydrogenase (LDH), which then enter the bloodstream (Hong et al., 2019). Higher plasma levels of these compounds indicate greater heat stress in cattle (Jiang et al., 2025).

The negative impacts of heat stress on dairy cattle when maintained above their comfort zone can be mitigated through nutritional strategies, such as feed supplementation. These include bypass protein, Ca-PUFA, and organic minerals. These supplements help by stimulating lipid metabolism, boosting immunity, preventing inflammation, and supporting thermoregulation, thereby reducing the adverse effects on the animal’s physiological condition. The nutritional modulation of dairy cow diets significantly impacts milk yield and the complex fatty acid profile of milk (Dhakal et al., 2024). This is vital for enhancing farm profitability and improving the nutritional quality of milk for consumers (Ahmed-Farid et al., 2021). Specifically, strategies such as supplementing with calcium salts of polyunsaturated fatty acids (PUFAs), protected proteins, and organic minerals have shown potential to influence ruminal biohydrogenation and nutrient partitioning, thereby directly affecting milk fat production and overall yield (Herrera et al., 2024). The goal of incorporating these dietary elements is to reduce the breakdown of beneficial nutrients in the rumen, increasing their availability for milk synthesis in the mammary glands (Lee et al., 2020; Mosoni et al., 2023). This approach employs advanced nutritional science to target metabolic pathways in dairy cows, enabling more efficient conversion of feed into high-quality milk components (Agle et al., 2020; Mosley et al., 2017). This study reviews current research on the effects of Ca-PUFA, protected proteins, and organic minerals on milk production and fatty acid composition, focusing on their mechanistic roles in modulating ruminal fermentation and systemic metabolism.

MATERIALS AND METHODS

Materials

The experimental animals used in this study were 32 Friesian Holstein (FH) dairy cows in the third lactation period, which were randomly distributed into predetermined treatment groups. The study was conducted over 16 weeks in an open housing system. The experimental animals were housed in individual pens measuring 1.5 x 2 m, each pen equipped with a treatment label. Feed was provided in limited quantities in accordance with standard requirements, and drinking water was provided ad libitum. EBI 25–TH Wireless Data Logger has been used in this research to record temperature and humidity in real time. The entire research procedure is visualized in Figure 1.

 

Treatment and housing microclimate

This study was conducted using a completely randomized design (CRD). It consisted of four treatments, each with four replicates. The four treatments were as follows, FS0: Control ration consisting of 60% forage (f) + 40% concentrate (ctr) ; FS1: 60% f + 40% ctr (97% ctr + 3% bypass protein); FS2: 60% f + 40% ctr (95% ctr + 3% bypass protein + 2% Ca-PUFA); FS3: 60% f + 40% ctr (93% ctr + 3% bypass protein + 2% Ca-PUFA + 2% organic minerals).

Temperature and humidity recordings were performed during the study. The average daytime temperature and humidity were 31°C and 89%, respectively. At night, they were lower, at 26°C and 78%, respectively.

Feed supplement preparation

The feed supplements in this study included bypass protein, Ca-PUFA, and organic minerals. Bypass protein was produced by mixing tannin with fishmeal. The process involves tannin binding to the amino and/or carboxyl groups of the protein (Tanuwiria et al., 2025).

Tannin-Protected Fish Meal Prepation, Fish meal was homogeneously sprayed with a tannin extract solution at 3% w/w of the fish meal and then dried in an oven at 6000C for three days. The nutrient composition of the tannin-protected fish meal was 17.09% water, 32.29% ash, 28.56% crude protein, 12.58% crude fat, 0.77% crude fiber, and 25.80% BETN.

The Ca-PUFA complex was produced via saponification using peanut oil and calcium hydroxide (Ca(OH)₂). As noted by Tanuwiria et al. (2025), the process involves hydrolyzing oil with a base to produce glycerol and fatty acid salts, in which the COOH groups of fatty acids bind to base cations. The Ca-PUFA was administered at 120 grams per fish daily. The PUFA from the peanut oil used consisted of 35% linoleic acid.

Organic minerals are produced by mixing zinc (Zn), copper (Cu), selenium (Se), and chromium (Cr) into corn and soybean media, which are then fermented with the help of Saccharomyces cerevisiae (SC) and/or Aspergillus oryzae (AO). The principle of producing organic Zn, organic Cu, organic Se, and organic Cr is to incorporate Zn, Cu, Se, and Cr into the proteins of the fungi Saccharomyces cerevisiae (SC) and/or Aspergillus oryzae (AO). The procedure for producing Zn-organic, Cu-organic, Se-organic, and Cr-organic compounds is as follows: the base substrate is a mixture of corn flour and soybean flour. The substrate is mixed with a standard solution (0.5% NH4NO3, 0.05% KCl, 0.05% MgSO4.7H2O, 0.001% FeSO4.7H2O, 0.0001% CuSO4.5H2O) and a solution of Zn, Cu, Cr, and Se at the specified dosage. The substrate is then sterilized at 121°C, 15 psi for 15 minutes. Next, it was inoculated with A. oryzae and S. cerevisiae cultures at a 2% (2 g per 100 g of substrate) inoculum. Incubation was carried out for four days at room temperature. The resulting product was dried in an oven at 60°C and ground. Bound minerals were quantified using standard procedures (Tanuwiria et al., 2025). The administration of organic Zn and organic Cu was 120 grams/animal/day, while organic Cr and organic Se were 10 g/animal/day.

Feed administration

Feed was provided twice daily at 07:00 and 15:00. The feed consisted of 6 kg of concentrate supplemented with feed supplements in the form of bypass protein, Ca-PUFA, and organic minerals according to the treatment, as well as forage equivalent to 10% of the dairy cow’s body weight.

Sample collection and analysis

Sampling was conducted twice, at the midpoint and end of the study, namely in the 5th, 10th and 16th weeks. Blood samples were obtained from the coccygeal vein using 3 mL tubes containing the anticoagulant Ethylenediaminetetraacetic Acid (EDTA).

Blood samples were analyzed using a hematology analyzer to determine erythrocyte, leukocyte, and differential counts. Stress biomarker parameters were obtained from blood plasma through centrifugation for 5 minutes at 5000 rpm. The concentration was measured with a spectrophotometer by mixing reagents and buffer solutions according to the Biolabo Kit analysis procedure, using a wavelength appropriate for the parameter (500–550 nm).

Data analysis

The data were analyzed using a completely randomized design variance analysis. Differences between treatments were determined with Duncan’s multiple range test. All analyses were performed with IBM SPSS 25 at a 5% significance level or 95% confidence level.

RESULTS AND DISCUSSION

Thermoregulation and hematological

The study’s Table 1 illustrates how adding the feed supplement to the diet influences thermoregulation and leukocyte differentiation, including neutrophils, lymphocytes, monocytes, eosinophils, and basophils.

The analysis of variance shows that feed supplements significantly affect leukocytes, neutrophils, lymphocytes, monocytes, eosinophils, and basophils (P<0.05). Duncan’s multiple range test was used to compare treatment means. According to Table 1, leukocyte levels decrease from FS0 to FS3, with values of 9.38 x10³/µL, 8.89 x10³/µL,

 

Table 1: Average thermoregulatory and hematological levels in lactating dairy cows with feed supplement administration of various treatments.

Parameter	Treatment
	FS0	FS1	FS2	FS3
Thermoregulation
Heart rate ( x minute-1)	78.15a	75.25ab	74.50b	73.76b
Respiratory rate ( x minute-1)	32.64a	30.83ab	29.28b	27.35b
Rectal temperature (0C)	39.05a	38.15a	38.05a	38.05a
ATPase Na+/K+ Transporting Subunit Alpha 1 (ng/dL)	3.04±0.02	2.83±0.05	2.26±0.06	1.75±0.03
Thyroxine (ng/dL)	1.04±0.03a	1.62±0.07b	1.93±0.04b	2.32±0.03c
Haematological
Red blood cells (x10⁶/µL)	5.87a	6.12b	6.15b	6.10b
Haemoglobin (g%)	7.15a	9.20b	10.15b	10.10b
Haematocrit	24.55a	27.41b	28.75b	30.34c
Leukocytes ( ×10³/µL )	9.38± 0.21c	8.89± 0.24b	8.63± 0.28b	8.68± 0.61a
Neutrophils (x 10²/µL)	7.01±0.07d	2.56± 0.18c	2.28± 0.24b	2.59±0.25a
Lymphocytes (x 10² /µL)	3.65± 0.11c	5.36± 0.07b	5.37± 0.04b	5.14± 0.90a
Monocytes (x 10²/µL)	0.23±0.04d	0.18± 0.04c	0.42± 0.06b	0.33±0.07a
Eosinophils (x 10²/µL)	0.27±0.01c	0.32±0.01b	0.23±0.01b	0.29±0.02a
Basophils (x 10³/µL)	0.10± 0.00c	0.01± 0.00b	0.02± 0.00ab	0.04± 0.01a

 

a,b,c,dDiDifferent superscripts on the same row indicate significant differences (P<0.05).

 

8.63 x10³/µL, and 8.68 x10³/µL, respectively. These levels remain within the normal range of 5.83 to 12.23 x10³/µL, as reported by Tanuwiria et al. (2022). The rise in leukocyte levels may be linked to increased neutrophil counts (especially in the FS0 group) or to changes in lymphocyte counts, since both cell types are predominant in the blood circulation.

Neutrophils can be a good and accurate indicator of ruminant stress. Neutrophils function to phagocytose and kill organisms (Jain et al., 2025). The average neutrophil count in experimental lactating dairy cows ranged from 2.28 to 7.01 x10³/µL (Table 1). The results of this study were generally within the normal range reported by Jiang et al. (2025), which was 1.7-6.0 x10³/µL, except for the FS0 group, which showed a slight increase.

Lymphocytes play an important and comprehensive role in the body’s defense system. The average lymphocyte count in this study ranged from 3.65 to 5.37 x10³/µL (Table 1). The average lymphocyte count was higher in the FS0 treatment group (4.65 × 10³/µL) than in the control group. This value is still within the range reported by Hong et al. (2019) and Guo et al. (2025), which is 1.8-8.1 x10³/µL.

Monocytes protect the body from invading organisms through phagocytosis (Goetz et al., 2022). The results of this study show that the average monocyte levels in lactating dairy cows range from 0.18 to 0.42 x10³/µL (Table 1). However, the average monocyte count was higher than the values reported by Rahayu et al. (2023) and Razzaghi et al. (2022), who reported a normal monocyte count of 0.3 x10³/µL in Friesian Holstein (FH) dairy cows. The high average monocyte count in this study is thought to be a response to heat stress. This is supported by D’Oliveira et al. (2021) and Guinard-Flament et al. (2024), who emphasize that animal stress can be assessed using monocyte counts and that stress factors can increase them.

One way to evaluate stress resistance is to administer multi-feed supplements and assess livestock’s response to feed additives that boost immunity. Eosinophils, basophils, and monocytes are appropriate parameters for this purpose. Eosinophils are produced in large numbers in patients with parasitic infections and migrate to tissues (Bach et al., 2005; Gobikrushanth et al., 2023; Chena et al., 2024). The average eosinophil count in this study ranged from 0.23 to 0.32 x10³/µL (Table 1). This value is slightly lower than that reported by Cavallini et al. (2025), who stated that the normal eosinophil count in Friesian-Holstein (FH) dairy cows ranges from 0.6 x10³/µL. However, the results of this study align with those of Michalowski (2005), who reported that the normal eosinophil count in cows ranges from 0-1.5 x10³/µL.

Basophils play a role in the development of hypersensitivity reactions and in the secretion of vasoactive mediators (Cuervo et al., 2025). This study showed that the average basophil count in lactating dairy cows ranged from 0.01 to 0.10 x10³/µL (Table 1). These results are in line with the view of Franzolin et al. (2010) that the normal basophil count in cows is 0.01-0.03 x10³/µL. Basophils are the myeloid cells with the lowest circulating concentrations and are commonly found in inflammatory and allergic conditions (Denton et al., 2022; Abouelezz et al., 2022).

Based on this study’s Findings, leukocyte counts and differentiation in experimental lactating dairy cows receiving feed supplements tended to be lower than in the control group. The FS3 treatment, which included 60% forage and 40% concentrate (comprising 93% concentrate, 3% bypass protein, 2% Ca-PUFA, and 2% organic minerals), resulted in the lowest average leukocyte differentiation profile (P<0.05) among all treatments. According to studies by Barłowska et al. (2023) and Puastuti et al. (2021), both genetic and environmental factors impact leukocyte levels and differentiation, with environmental influences including infection and feed.

Leukocyte formation and differentiation depend on amino acids for protein synthesis. Bypass protein supplementation provides many amino acids crucial for supporting the immune response during heat stress in dairy cows, as these amino acids improve immune function and gluconeogenesis (Tanuwiria et al., 2022; Razzaghi et al., 2022). Moreover, amino acids boost protein metabolism, help alleviate heat stress, and increase blood concentrations, especially leukocyte counts, to within normal levels (Mosoni et al., 2023; Michalowski, 2005).

Research by Wijayanti et al. (2022) and Theodorou et al. (1994) confirms that PUFA effectively mitigates heat stress, enhancing FH cattle performance and immune health during hot conditions. Additionally, PUFAs can elevate leukocyte levels and support their differentiation within normal ranges. They also influence immune and inflammatory responses, playing a crucial role in cellular metabolism and leukocyte function (Mosley et al., 2017; Barloska et al., 2023).

Administering organic minerals can enhance enzyme and hormone activity and support metabolic processes in the body (Abouelezz et al., 2022; Ahmed-Farid et al., 2021). Essential organic minerals such as chromium (Cr), copper (Cu), selenium (Se), and zinc (Zn) are vital for dairy cow health. Cr supplementation can increase milk production, boost immune function, and enhance overall health, especially in stressed cows and during early lactation (Chena et al., 2024). Copper (Cu) is involved in cellular energy metabolism, nerve impulse transmission, and the health of the cardiovascular and immune systems (Denton et al., 2022). Its biological roles include ceruloplasmin, superoxide dismutase (SOD), lysine oxidase, and cytochrome oxidase (Jiang et al., 2025). Moreover, Cu improves immunity and fertility (D’Oliveira et al., 2021). Selenium (Se) functions as an antioxidant within selenium metabolism to maintain homeostasis (Dhakal et al., 2024), reducing cell damage caused by heat stress. Se also modulates immune responses in livestock by activating neutrophil phagocytosis, increasing antibody production, and promoting lymphocyte proliferation (Tanuwiria et al., 2022; Jiang et al., 2025). Zinc (Zn) has anti-inflammatory properties and participates in many biological functions (Muslim et al., 2014; Mosoni et al., 2023).

The decrease in ATP1A1 levels or expression in dairy cows that are not experiencing heat stress reflects the reduced cellular need to maintain ionic homeostasis previously disrupted during heat stress. Under thermoneutral conditions, body temperature, electrolyte balance, and acid-base status return to stability, resulting in decreased Na⁺ and K⁺ transport activity across cell membranes (Mosoni et al., 2023). Consequently, the energy requirement for operating the Na⁺/K⁺-ATPase pump is reduced, and ATP1A1 expression is re-regulated toward basal levels. This mechanism represents a physiological adaptation aimed at increasing the efficiency of cellular energy use when environmental stress has subsided and reducing unnecessary metabolic load.

Stress biomarkers for inflammation

The effects of feed supplement administration in rations on stress biomarkers, including alkaline phosphatase, creatinine, creatine kinase, Gamma Glutamyl Transpeptidase (γ-GT), and Lactate Dehydrogenase (LDH), are shown in Table 2, based on the study.

The variance analysis showed that administering feed supplements had a significant effect (P<0.05) alkaline phosphatase, creatinine, creatine kinase, gamma glutamyl transpeptidase (γ-GT), and lactate dehydrogenase (LDH). Furthermore, Duncan’s multiple range test was conducted to assess differences in means among treatments. As shown in Table 2, the alkaline phosphatase (ALP) parameter differed significantly (P<0.05) across treatments. Based on the results of the study, lactating dairy cows given the FS0 treatment had higher averages than FS1 to FS3, which were 43.33 U/L, 37.58 U/L, 37.31 U/L, and 34.52 U/L, respectively. Previous research by Lee et al. (2020) reported that ALP levels in dairy cows decreased with increasing parity.

Table 2 presents the effect of feed supplement administration on creatinine levels in lactating dairy cows. The study shows that cows in FS0 had significantly higher creatinine levels (123.41 µmol/L, P<0.05) than those in FS1 to FS3, which recorded 109.46 µmol/L, 99.38 µmol/L, and 96.52 µmol/L, respectively. These values fall within the typical range reported by Ahmed-Farid et al. (2021), 97.5-111 µmol/L. Elevated blood creatinine levels can indicate kidney issues such as glomerular filtration problems, acute tubular necrosis, dehydration, or renal failure (Gobikrushanth et al., 2023; Herrera et al., 2024).

 

Table 2: Levels of stress biomarkers in lactating dairy cows with feed supplement administration.

Parameter	Treatment
	FS0	FS1	FS2	FS3
Malondialdehyde (MDA) (ng/dL)	3.72±0.06a	2.93±0.17a	1.05±0.08b	0.95±0.07b
Total Antioxidant (ng/dL)	0.13±0.04a	0.22±0.05b	0.46±0.03c	0.93±0.03d
Interleukin-6 (ng/dL)	2.76±0.07a	2.62±0.04a	2.04±0.04b	1.73±0.02c
Creatinine (µmol/L)	123.41±3.22d	109.46±4.03c	99.38±3.33b	96.52±3.11a
Creatine Kinase (U/L)	4.33±0.06d	2.22±0.02c	1.85± 0.11b	0.36±0.02a
Lactate dehydrogenase (LDH) (U/L)	18.53±1.11c	11.05±0.22b	8.84±0.44ab	7.01±0.42a
Gamma-Glutamyl Transpeptidase (U/L)	16.63±1.04d	14.34±1.06c	11.63±1.01 b	8.44±0.11a
Alkaline Phosphatase (U/L)	43.33±1.46c	37.58±1.42b	37.31±1.42b	34.52±1.24a

 

a,b,c,d Different superscripts on the same row indices significant differences (P<0.05)

 

Table 2 shows the effect of feed supplement administration on creatine kinase in lactating dairy cows. Each treatment differs significantly (P<0.05). Based on the results of the study, the average creatine kinase in lactating dairy cows given the FS0 treatment was higher (4.33 U/L) than in those given FS1 (2.22 U/L), FS2 (1.85 U/L), or FS3 (0.36 U/L). According to Michalowski (2005) and Chena et al. (2024), heat stress causes muscle metabolism to provide energy by breaking down creatine phosphate into creatinine via the enzyme creatine kinase. This indicates that high plasma levels of creatinine and creatine indicate that the cattle are experiencing heat stress.

Gamma-glutamyl transpeptidase (γ-GT) is an enzyme prevalent in cardiac cells. Homeostatic mechanisms related to cardiac function (Gobikrushanth et al., 2023; Denton et al., 2022), especially those governing energy supply (Razzaghi et al., 2022), can lead to increased cardiac cell death (necrosis). Past research indicates that necrosis promotes metabolite leakage into the bloodstream (Puastuti et al., 2021; Rahayu et al., 2023; Mosoni et al., 2023). As shown in Table 2, γ-GT levels were significantly higher (P<0.05) in lactating dairy cows fed only FS0 (control diet) than in those fed FS1, FS2, and FS3. The findings reveal notable differences (P<0.05) in γ-GT levels across treatments. Levels declined from FS0 to FS3, measuring 16.63 U/L, 14.34 U/L, 11.63 U/L, and 8.44 U/L, respectively. Overall, the FS3 group exhibited the lowest average γ-GT level across treatments.

Table 2 shows the effect of feed supplement administration on LDH levels. According to the table, LDH levels in FS0 were significantly higher (P<0.05) at 18.53 U/L than in FS1-FS3, which received the feed supplement treatment. Their LDH levels were 11.05 U/L, 8.84 U/L, and 7.01 U/L, respectively. Elevated blood LDH levels can indicate damage to cells, muscle membranes, and tissues (Franzolin et al., 2010; Bach et al., 2005) and also serve as an early blood marker for heat stress.

This study showed that the levels of certain metabolic compounds in the blood such as Alkaline Phosphatase (ALP), creatinine, creatine kinase, Gamma Glutamyl Transpeptidase (γ-GT), and Lactate Dehydrogenase (LDH) which serve as stress indicators in experimental lactating dairy cows receiving feed supplements, tended to be lower on average than in the control group. The FS3 treatment, composed of 60% forage and 40% concentrate (including 93% concentrate, 3% bypass protein, 2% Ca-PUFA, and 2% organic minerals), showed the lowest stress levels (P<0.05) among all treatments.

Milk production and fat profile

Table 3 shows milk production and fat profile from dairy cows raised in lowland areas with feed supplement administration. Current research results indicate that administering FS2 and FS3 significantly increases average milk production (P<0.05) compared to the experimental group of cows without feed supplements and those receiving FS1. The content of long-chain polyunsaturated fatty acids also appears to be higher in cows that received rations supplemented with FS2 and FS3. In contrast, the saturated fatty acid content was lower.

Previous research indicates that concentrates rich in crude protein and Ca-FA can markedly increase intake of dry matter, protein, fat, and energy, thereby boosting milk production (Puastuti et al., 2021). Additional energy, particularly from fatty acids, can increase milk fat and protein yields (Mosley et al., 2017).

Adding Ca-PUFA to dairy cow diets has been shown to alter macromineral and metabolic marker levels in the bloodstream, potentially reducing postpartum health issues and improving reproductive efficiency (Gobikrushanth et al., 2023). Additionally, increased nutrient uptake by the mammary gland from Ca-PUFA supplementation could explain the rise in milk fat levels by altering nutrient utilization (Guinard-Flament et al., 2024). Calcium is

 

Table 3: Milk production and fat profile of dairy cows in lowland areas with feed supplement addition.

Milk production and fat profile	Feed supplement
	FS0	FS1	FS2	FS3
Average daily milk production (L)	13.5±1.31a	13.7±0.67a	14.87±1.65b	16.85±2.03c
Fatty acids (%)
C4:0	3.87±0.61a	2.92±0.04a	2.36±0.16c	2.05±0.03d
C6:0	2.64±0.01a	2.56±0.11a	2.15±0.11ab	2.08±0.04b
C8:0	1.62±0.06a	1.64±0.07a	1.37±0.01ab	1.28±0.03b
C10:	4.28±0.84a	3.94±0.64a	3.32±0.21ab	3.11±0.02b
C12:0	4.44±1.04a	4.39±0.51a	3.85±0.11ab	3.42±0.03b
C14:0	11.12±1.53a	11.02±1.03a	10.44±1.43a	10.07±1.95b
C16:0	28.71±3.034a	28.74±3.05a	28.28±1.53ab	26.13±2.42b
C18:0	12.21±2.64a	12.19±1.51a	12.16±1.07ab	11.17±1.05b
18:1 cis-9	24.34±3.02a	24.38±2.55a	24:56±2.35a	25.55±3.17b
18:2 cis-9, cis-12	3.57±0.31a	3.57±0.333a	3.92±0.17a	4.48±0.09b
total n-6	3.83±0.53a	3.88±0.42a	4.08±0.44a	5.17±0.81b
total n-3	0.36±0.07a	0.55±0.02a	1.16±0.02b	1.38±0.06b
SFA	70.25±3.70a	69.77±2.64a	67.25±3.05b	66.17±3.45b
MUFA	27.08±1.48a	27.26±1.64a	28.57±1.63b	29.17±1.57c
PUFA	4.08±0.72a	4.41±0.66a	4.53±0.15a	5.87±0.85b
Total fat (g.100g -1)	5.15±0.92a	5.26±0.47	5.41±1.63a	5.64±0.44b

 

a,b,c,d Different superscripts on the same row indicate significant differences (P<0.05). SFA: saturated fatty acids; MUFA: monounsaturated fatty acids; PUFA: polyunsaturated fatty acids.

 

Organic minerals play a crucial role in shaping the fatty acid profile of dairy cow milk, influencing milk fat synthesis and desaturation (Razzaghi et al., 2022). For instance, certain organic trace minerals, such as zinc and selenium, serve as cofactors for enzymes involved in fatty acid metabolism, potentially affecting the concentration of beneficial unsaturated fatty acids in milk (Barłowska et al., 2023). This suggests that targeted supplementation with organic minerals could be an effective strategy for dairy farmers seeking to enhance the nutritional value and functional qualities of milk (Puastuti et al., 2021; Cavallini et al., 2025).

CONCLUSION

This study concluded that providing feed supplements effectively reduced the risk of heat stress in dairy cattle in Indonesia’s lowlands. Physiological indicators like thermoregulation and hematology remained normal or improved, and free radical activity decreased even under heat stress. The most effective treatment was FS3, which involved lactating dairy cows fed a diet comprising 60% forage and 40% concentrate, including 93% concentrate, 3% bypass protein, 2% Ca-PUFA, and 2% organic minerals.

ACKNOWLEDGMENT

The author wishes to thank everyone who supported and helped with this research, especially the Ruminant Nutrition and Animal Feed Chemistry laboratory assistants and the Animal Physiology and Biochemistry laboratory at Padjadjaran University.

NOVELTY STATEMENT

This study’s key innovation is the use of a multi-component feed supplement that combines calcium-protecting polyunsaturated fatty acids (Ca-PUFA), organic oils, and bypass proteins as a comprehensive nutritional strategy. This combination is believed to act synergistically to alleviate heat stress in dairy cattle raised in lowland tropical regions, particularly in Indonesia, a novel application not previously reported. The approach broadens current knowledge by examining how these feed components interact to influence thermophysiological responses focusing on specific thermoregulatory markers such as ATPase Na+/K+ Transporting Subunit Alpha 1 and interleukin-6, which indicate free radical activity and productive performance. Overall, it offers a sustainable approach to enhancing the resilience and productivity of dairy cattle under chronic heat stress.

AUTHORS CONTRIBUTION

The authors listed in this article have contributed equally to its preparation and writing. UHT, AM: Planned and designed the research, supervised implementation, guided sample analysis, analyzed and interpreted data, and wrote, edited, and finalized the article. BKM, AH: Conducted research in the barn, analyzed samples, performed statistical analysis, and authored the article.

Ethical approval

This experiment was designed and conducted with the understanding that no experimental techniques violated animal experimentation ethics rules, as approved by the Research Supervisory Agency, DP Indonesia, under number 183/Ret/rt.23/2025.

Generate AI and AI-assisted technology statement

The authors declare that they did not utilize any technological assistance in the form of AI software or similar tools in drafting and writing this article or in the data analysis contained herein.

Conflict of interest

The authors have declared no conflict of interest. All scientific information and research data published in this article are stated to have no conflict of interest whatsoever with any party, whether in the form of patents, financing, or rights related to them.

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