Animal Science

Energy-mineral supplementation during the gestation-lactation transition period in Holstein cows

0000-0001-5546-1259Luis Rodrigo Balarezo-Urresta¹  , 0000-0001-6364-3546Ernesto Noval-Artiles²  , 0000-0002-2966-7824Juan Ramón García-Díaz^{2  *} 

¹Universidad Politécnica del Carchi. Facultad de Industrias Agropecuarias y Ciencias Ambientales. Tulcán, Proyecto de mejoramiento de la competitividad láctea en la provincia del Carchi. Escuela de Desarrollo Integral Agropecuario, Carchi, Ecuador.

²Universidad Central “Marta Abreu” de Las Villas. Facultad de Ciencias Agropecuarias, Departamento de Medicina veterinaria y Zootecnia, Carretera a Camajuaní Km. 5 ½. Santa Clara. CP 54830, Santa Clara, Villa Clara, Cuba.

Abstract

In order to determine the effect of energy-mineral supplementation during the gestation-lactation transition period on the metabolic profile and reproductive indicators of Holstein cows,a total of 32 animals, between the third and fourth lactation, and body condition ≥ 3.5 were selected. The experiment was performed in Carchi province, Ecuador, during the rainy season. Two groups of 16 animals each were randomly formed. The same foods were supplied to all, and those in the treated group were supplemented with corn, in an amount of 1.0 kg DM, 30 days before parturition, and 2.0 kg DM at the beginning of lactation until 60 days postpartum, plus parenteral supplementation of 20 mL of kyrofosfan.The hematochemical parameters and reproductive indicators were determined, which were compared by t- Student test for independent samples. The contributions of Cu and Zn did not cover the requirements in both study groups. The metabolizable energy was deficient in the animals that were not supplemented with corn. In the cows from the treated group, the concentrations in blood serum of P, Cu, Zn and cholesterol, and the body condition were higher (P <0.05) at 30 and 60 days postpartum. Total proteins increased (P <0.05) at 60 days postpartum, while betehydroxybutyrate decreased (P <0.01) at 30 days postpartum.The supplementation decreased (P <0.05) the parturition-first service interval, the services per gestation (P <0.001) and the parturition-conception and parturition-parturition intervals. It is concluded that energy-mineral supplementation improves the metabolic profile and reproductive indicators.

Key words: energy; protein; deficit; metabolic profile; reproductive indicators

In tropical regions, most grasses do not meet the mineral requirements of grazing dairy cows, so there are marked deficiencies that are associated with reproductive problems (^{McDowell and Arthington 2005} and ^{García et al. 2010}). In catlle herds, with an apparently adequate diet, alterations in protein, mineral and energy metabolism were diagnosed, with a manifest deterioration in reproductive capacity (^{García et al. 2011}).

In catlle herds of the Andean region from Ecuador, more than 30 % and 65 % of cows were diagnosed with high concentrations in blood serum of urea nitrogen (BUN) and betahydroxybutyrate (B-OH), respectively, and more than 60 % of females with hypophosphataemia, hypocupremia and hypocinemia. All these nutritional and metabolic alterations were associated with the deterioration of reproductive indicators (^{Balarezo et al. 2016}).

The energy in food is the limiting factor of reproduction for cows in the gestation-lactation transition period, which includes three weeks before and after parturition (^{Castro et al. 2012}). However, there are criteria that suggest that the first weeks of the dry season should also be analyzed (^{Esposito et al. 2014}).

The energy deficit causes an increase in B-OH, as a consequence of the mobilization of fats for obtaining energy, and decreases body condition, mainly during the transition period of the dairy cow. Consequently, energy deficiency affects the restart of postpartum ovarian activity (^{LeBlanc 2010}) and uterine involution (^{Balarezo et al. 2018}).

In addition to the previous, the energy deficit in the ration causes the negative energy balance (NEB) in the cows, which causes the disorder of the functions of the hypothalamic-hypophysis -ovary axis; and it affects the secretion, amplitude, and frequency of GnRH and LH pulses, which can cause reproductive problems (^{Walsh et al. 2007}).

The objective of this study was to determine the effect of energy-mineral supplementation, fitted to the production conditions of the Andean region from Ecuador, on the metabolic profile and reproductive indicators of Holstein cows during the gestation-lactation transition period.

Materials and Methods

Location. The research was developed from August 2016 to July 2017, in Tufiño parish, Tulcán canton, El Carchi province. This region is located between the 1º12ʼand 43" North Latitude and 78º 33ʼ 12” West Longitude, an altitude of 2990 and 3450 m o.s.l. The soil is of the Andisol order, with an effective depth of 70 cm. The relief is undulating, with a slope percentage that fluctuated between 10 and 20 % (^{Balarezo et al. 2016}), determined with GPS map 60 CSx.

The rainfalls ranges between 1 000 and 1250 mm in the rainy season (RS), from October to April, and from 700 to 850 mm, in the dry season (DS), from May to September. The average temperature fluctuates between 6 and 11°C, with minimums and maximums of 2 and 15°C, respectively.

Experimental design. A total of 32 Holstein cows were selected, between the third and fourth lactation, with an age between 4 and 8 years, and body condition (BC) ≥ 3.5. They were in rationed grazing by electric fence for 18 h. The bromatological composition of the supplied food and the intake is showed in table 1.

Table 1 Bromatological composition and intake of the supplied food to the studied animals 

Foods	DMI (kg)	DM (%)	ME (MJ kg DM^-1)	CP (%)	Ca	P	Cu	Zn
					%		(mg kg DM^-1)
Zea mays	2.00	87.00	13.35	8.1	0.30	0.29	2.05	23.30
Lolium perenne	4.77	14.10	11.51	21.35	0.27	0.59	5.00	10.00
Cenchrus clandestinun	2.12	14.00	8.49	20.37	0.26	0.35	13.90	59.50
Holcus lanatus	0.53	15.80	8.91	17.00	0.42	0.59	3.70	21.70
Trifolium repens	0.42	13.40	9.24	28.00	1.25	0.52	8.80	31.00
Plantago sativa	2.75	10.60	9.41	23.80	1.16	0.44	12.4	21.00
CALFOSAL	0.15	98.00	------	-------	12.00	6.00	2.90	1.60

DMI:Dry matter intake, DM:dry matter, ME: metabolizable energy, CP: crude protein

Calfosal (multisalmin SA, Ecuador) was supplied, which has a Ca: P ratio of 2.25: 1 (table 2). It was orally applied, to guarantee between 100 to 150 g animal^-1 d^-1 of availability. Two groups of 16 animals each were randomly established . One group was used as a control and the other as a treated.

Table 2 Chemical composition of calfosal used as oral supplementation 

Mineral	Concentration, %	Mineral	Concentration, %
P	6.00	Cu	0.29
Ca	12.00	Co	0.002
Mg	0.30	S	0.16
Na	17.00	Cl	27.00
K	0.005	Fe	0.15
Mg	0.30	Se	0.0003
Zn	0.16	Mn	0.004
I	0.02

The animals from the treated group were given corn, in an amount of 1.0 kg DM, 30 d before parturition , and 2.0 kg DM at the beginning of lactation until 60 d postpartum (DPP). In addition, 20 mL of kirofosfan (Kirovet SA, Colombia) was parenterally applied, at 30 days before parturition (DBP), at parturition, and at 30 DPP. It contains sodium toldimfos (200 mg), zinc sulfate (1.1 mg), sodium selenite (0.33 mg), nicotinic acid (5 mg), potassium iodide (20 mg), manganese sulfate (1 mg) and 40 mg. copperxilin (Brouwer, Argentina), which contains copper glycinate (10 mg).

Taking blood samples. For the biochemical analyzes, 10 mL of blood was extracted by coccygeal venipuncture with vacutainer tubes, without anticoagulant. It was waited 24 h for the spontaneous retraction of the coagulum, it was centrifuged at 2,500 g for 10 min. and blood serum was obtained, which was frozen at -10°C until the analysis.

Determination of hematochemical parameters.The hemochemical indicators were determined in a Star Fax 3300 equipment (Aznar Diagnóstica, USA), using commercial kits, according to the manufacturer's procedures (table 3). All the analyzes were carried out in the diagnostic laboratory, Carlos Martínez Hoyos veterinary clinic, from Universidad de Nariño, Colombia.

Table 3 Hematochemical indicators evaluated in the researched cows 

Profile	Variables	Method
Mineral	Ca, Mg, Cu, Zn	EEA (Miles et al. 2001)
	Phosphorus	Phosphomolybdate. Ultra Violet (UV)
Hematochemical	Total proteins	Biuret. Colorimetric
	Cholesterol	CHOD-POD. Enzymatic colorimetric
	β-hydroxybutyrate (B-OH)	RANBUT D-3. Enzyme kinetic

Body condition. The BC was estimated by physical examination of the animals, which included inspection and palpation. It was classified on a scale of 1-5 points and divisions of 0.25 between them, according to the methodology proposed by ^{Rodenburg (2004)}.

Methodology for heat and artificial insemination detection. Heat detection was performed from 5-9 a.m. and from 3-7 p.m., by a trained man. As an auxiliary method, paint was used on the base of the cows' tails. An experienced technician carried out the insemination by depositing the semen in the body of the uterus, with a technical efficiency between 60 and 65 % in the last four years. Semen from proven fertility bulls was used, content in straws of 0.25 and concentrations of 32 million per dose.

Analysis of reproductive indicators. The parturition-first service intervals, parturition-conception interval, parturition-parturition interval, percentage of conceptions at first service and services per conceptions were determined. It was proceeded from individual records, according to the methodologies described by ^{Brito et al. (2010)}.

Statistical processing. The biochemical and reproductive indicators were compared between the control and treated groups by means of the t- Student test for independent samples. The statistical package StatgraphisCenturion Ver. XV.II (^{StatPoint Technologies 2010}) was used in these processes.

Results and Discussion

Table 4 shows the nutrient balance. At 35 DPP, in the treated group, the incorporation of corn and the supplementation of mineral salt orally satisfied the nutrients of metabolizable energy and Ca, respectively. However, the control group maintained the energy imbalance.

Table 4 Nutrient balance of the treated group at 35 DPP 

Food	ME (MJ d^-1)	CP	Ca	P	Cu	Zn
		g d^-1			mg d^-1
Control group
Total contribution	140.92	3 005.17	87.80	79.40	114.62	397.11
Requirement	145.78	1 789.00	87.09	47.01	140.00	561.66
Difference	-4.86	1 216.17	0.71	32.39	-25.37	-164.55
Treated group
Total contribution	162.48	3 071.00	94.11	83.14	113.00	421.01
Requirement	153.98	1 926.00	93.3	49.67	147.50	615.00
Difference	8.80	1 144.99	0.81	33.47	-34.455	-193.99

In both groups, the total CP contributions were higher than the requirements of these animals, and its concentration was in accordance with the estimated DM intake, which was 21.38 % in the control group and 19.37 % in the treated group; and 14.05 kg DM^-1 and 15.85 kg DM^-1, respectively. In the first case, the lower DMI could be due to excess protein, and the imbalance between this one and the energy (^{NRC 2001}).

The excess of CP in lactating animals, between 20 and 23 %, negatively affects the parturition-conception interval, decreases the pH in the uterus and increases the concentration of urea and P₄ in blood in cows in early lactation (^{NRC 2001} and ^{Castro et al. 2012}).

In both groups, the excess of N caused an energy expenditure to be able to eliminate it, increasing the open days, services per conception and interval between parturitions(^{David et al. 2018}). However, energy supplementation facilitated a better energy-protein ratio (table 4).

The concentrations of Ca and Mg in blood serum did not differ at any time (P> 0.05) in the supplemented cows with respect to the control during the gestation-lactation transition period (table 5).

Table 5 Concentration of minerals in blood serum in the gestation-lactation transition period 

Variables	Moment	Groups		±SE
		Control	Treated
Ca, mmol L^-1	30 DBP	3.68	3.43	0.20
	30 DPP	2.86	2.99	0.22
	60 DPP	3.61	3.80	0.16
P, mmol L^-1	30 DBP	1.75	1.79	0.08
	30 DPP	1.68	2.18	0.11**
	60 DPP	1.69	2.16	0.08**
Mg, mmol L^-1	30 DBP	1.25	1.20	0.06
	30 DPP	1.42	1.44	0.0767
	60 DPP	1.36	1.53	0.10
Cu, µmol L^-1	30 DBP	11.76	11.65	0.14
	30 DPP	11.00	12.07	0.21***
	60 DPP	11.16	12.62	1.61***
Zn, µmol L^-1	30 DBP	12.37	12.16	0.28
	30 DPP	11.98	13.50	0.20***
	60 DPP	12.41	14.33	0.21***

** P < 0.01, *** P < 0.001 in rows indicate significant differences between the the values shown in each of them (t-Student).

DBP: days before parturition

DPP: days postpartum

The Ca for the two groups was always above the critical limit (CL) of deficiency established by ^{McDowell and Arthington (2005)}. This agrees with the contribution of this mineral in the supplied food, which covered their requirements. Also, the dairy cow performs a strong action on Ca homeostasis, with the aim of preventing its loss, especially in highly productive animals or during prepartum (^{NRC 2001}).

In both groups, the Ca concentrations in blood serum decreased at 30 d of lactation with respect to the evaluation performed at 30 DBP. This was influenced by the high yield in dairy production and the start of production activity. However, as the Ca requirements are covered, the cow can support metabolic activity during the evaluated period.

In the research 88.8 % of the grasses samples showed deficient Mg values, according to the CLs suggested by ^{McDowell and Arthington (2005)}. However, the supplementation with Calfosal substitute the nutritional requirements of this macroelement, which is corroborated in its concentrations in blood serum in both groups (table 5), although they did not differ (P> 0.05). This may be mainly due to an adequate ruminal pH and the appropriate absorption of this mineral (^{NRC 2001}) from the gastrointestinal system (^{Schonewille 2013}).

The supplementation increased (P <0.01) the P concentrations in blood serum, at 30 and 60 DPP from 1.68 to 2.18 and 1.69 to 2.16 mmol/L, respectively (table 5), which must have decreased the losses of this macroelement due to its use in dairy production, and must have favored postpartum reproductive activity. For this reason, it is almost mandatory to supplement good quality minerals by different ways, including soil fertility enhancing procedures (^{McDowell and Arthington 2005}).

In the treated group, cupremia and cinchemia were higher (P <0.001) at 30 and 60 DPP with respect to the control group (table 5). In the latter, at all evaluation moments, they were lower than the deficiency CL established by ^{McDowell and Arthington (2005)}, consequently with the deficit of these microelements in the nutrient balance.

These results corroborate the ^{Noval et al. (2016)} reports, who refers that parenteral supplementation of 50 mg of Cu, 100 mg of Zn and 50 mg of Mn increased the cupremia and cinchemia, from five and 15 days after treatment, respectively. While, blood concentrations of both minerals become deficient at 60 d.

Under this research conditions, the P supplied in the diet covered the requirements of the dairy cows, but there were deficiencies of the macroelement in the blood serum of the animals, which could be caused by the excesses of CP and Mg in the diet.

The excessive contributions of CP and Mg originate in the rumen the formation of magnesium sulfate, which reduces the absorption of P in the small intestine (^{Djokovic et al. 2014}), and iron sulfide, which reduces the absorption of Cu at ruminal level (^{Arthington and Brown 2005}). In addition, in the soil there are excesses of S and Fe (^{Balarezo et al. 2016}) that condition the deficiency of Cu and Zn in the blood serum of cows (^{McDowell and Arthington 2005}).

These aspects show that altought P, Cu and Zn are supplemented orally, there were deficiencies of these minerals or, at least, this way will not be the most effective for supplementation. This justifies the strategy of oral and parenteral mineral supplementation used in this research, which increased the concentrations of P, Cu and Zn in blood serum (table 5).

These results agree with those obtained by ^{García et al. (2012}) and ^{Noval et al. (2016)}, who with the supplementation of other mineral sources, different schemes and parenterally, obtained favorable results in reproductive and economic efficiency.

The injectable minerals avoid interferences and antagonisms in their absorption and metabolism (^{García et al. 2012}). However, oral supplementation is very important for microorganisms and rumen bioactivity (^{McDowell and Arthington 2005} and ^{Djokovic et al. 2014}). The combination of both ways for the supplementation of minerals constitutes one of the scientific and practical conributions of this research.

Table 6 shows the concentrations of the energy and protein profile indicators of the cows under study. In supplemented cows, B-OH concentrations decreased (P <0.01) at 30 DPP. The BC and blood cholesterol concentrations were higher (P <0.05) in supplemented cows at 30 and 60 DPP. Also in them, total proteins (TP) increased (P <0.05).

Table 6 Levels of the energy and protein profile indicators in the cows under study during the gestation-lactation transition period. 

Variables	Moment	Groups		±SE
		Control	Treated
B-OH, mmol L^-1	30 DBP	0.51	0.47	0.04
	30 DPP	0.93	0.57	0.06**
	60 DPP	0.87	0.65	0.05
BC, U^-1	30 DBP	3.50	3.56	20.08
	30 DPP	2.61	2.93	0.08**
	60 DPP	3.21	3.50	0.07*
Cholesterol, mmol L^-1	30 DBP	3.48	3.28	0.11
	30 DPP	2.72	3.02	0.11*67
	60 DPP	3.42	3.80	0.08*
TP, g L^-1	30 DBP	91.31	92.06	0.62
	30 DPP	83.75	85.93	2.58
	60 DPP	88.37	94.37	1.61*

* P < 0.05, ** P < 0.01 in rows indicate significant differences between the the values shown in each of them (t-Student)

DBP: days before parturition

DPP: days postpartum

The decrease in B-OH is due to that in the animals supplemented with corn there was less fat mobilization to compensate the energy deficit. Under these conditions, the concentrations of non-esterified fatty acid in the follicular fluid decrease. In this medium, the oocytes have higher quality and competence and, consequently, higher fertility (^{Aardema et al. 2019}).

The improvement in energy metabolism indicators may be due to supplementation with corn, which increased body energy reserves and BC at 30 and 60 DPP.

These results are in contrast to those stated by other authors, who show that the peak of DM intake occurs at 10-14 weeks postpartum, but that of lactation occurs at the eighth week. Therefore, the NEB should be extended for more than 30 DPP (^{Macrae 2019}). The lack of correspondence can be explained because this experiment was performed with cows that had a BC ≥ 3.5 and controlled research conditions. The latter included a balanced ration and quality grasses , which allowed them to have a NEB of shorter duration.

The improvement in the energy status of the supplemented animals may be due to the increase in cupremia and cinchemia in them.The Cu is a co-factor of the terminal enzyme of the respiratory chain, and cytochrome-c-oxidase catalyzes the transfer of four electrons to O₂ to form two molecules of water and ATP, in addition to participating in obtaining energy (^{Gebhard et al. 2001}).

The Zn is involved in energy metabolism because the secretion of pyruvate kinase in the β cells of the pancreas is dependent on this microelement. This enzyme participates in glycolysis, so it is assumed that there is a compromise of the energy metabolism of the animal during Zn deficiency (^{Evans and Henshaw 2008}).

In cows supplemented with 50 mg of Cu parenterally, three applications every 60 days, increased (P <0.05) the BC (^{García et al. 2007}). This treatment in fattening bulls increased (P <0.05) live weight (LW) and their mean daily gain (MDG) (^{García et al. 2017}).The injectable Cu supplementation, alone or in combination with Zn and Mn, increased (P <0.05) the serum levels of Cu and Zn and MDG in calves (^{Noval et al. 2016}).These results corroborate the importance of these microelements to improve LW and MDG, both related to energy metabolism.

In the treated group, cholesterol values were higher (P <0.05) at 30 and 60 DPP (table 6), which corroborates the results obtained by ^{Moyano and Rodríguez (2014)}. The increase in cholesterolemia is associated with a better adaptation of the animals of this group to NEB, as a result of the supplementation with corn, the decrease in B-OH concentrations and the increases in cinchemia.The Zn is necessary for the synthesis of cholesterol, which is in turn essential for the synthesis of steroids, E₂, P₄ and T₄, which favor the adequate presentation of estrous cycles (^{Omur et al. 2016}).

Total proteins increased in supplemented animals at 60 DPP (table 6), possibly because the magnitude of NEB decreased and because of the beneficial actions in the protein metabolism of minerals, especially Cu (^{García et al. 2017}).

The main reproductive indicators (table 7) in the control group had a marked deterioration, according to the reference parameters of dairy cattle in optimal production conditions (^{Brito et al. 2010}). However, the mineral energy supplementation improved the indicators with respect to the control.

Table 7 Reproductive indicators of the study groups during the evaluated period 

Indicators	Groups		±SE
	Control	Treated
Parturition- first service interval, d	126.93	99.43	5.67*
Parturition -conception interval, d	181.31	138.62	9.49**
Parturition-parturition interval, d	463.31	420.62	9.23**
Service per conception, U	2.43	1.93	0.17*
Proportion of conceptions at first service	0.4115	0.5881	0.17

* P < 0.05, ** P < 0.01 in rows indicate significant differences between the the values shown in each of them (t-Student)

There are several mechanisms during pre parturation and post parturition that affect the fertility of the dairy cow. One of the main is NEB, which affects follicular growth and estradiol production, possibly due to decreased levels of insulin, insulin growth factor, and HL pulses. The loss of BC after parturition increases the percentages of cows that do not cycle at the end of the voluntary waiting period (^{Carvalho et al. 2014}).

The decrease (P <0.05) of the reproductive indicators in the supplemented cows is due to the supplementation increased the concentrations of P, Cu and Zn, which play important roles in the reproductive performance of the dairy cow. In Cuba, parenteral Cu supplementation in the gestation-lactation transition period favored reproductive performance (^{García et al. 2012}).

The increase in cupremia and cinchemia favor the production of E₂ and P₄ by luteal cells and, consequently, postpartum reproductive performance. In addition, Mn supplementation favors the synthesis of cholesterol, and this of the mentioned hormones (^{Griffiths et al. 2007}).

Also the energy supplementation of cows with the use of corn reduced the NEB and with it, the mobilization of fat and the B-OH. These circumstances caused an increase in BC and cholesterol in the supplemented cows.

The increase in cholesterol is associated with higher synthesis of E₂ and P₄, which promote a better uterine environment. This favors the maternal recognition of the pregnancy, the implantation of the embryo and the development of the pregnancy (^{Quintela et al. 2008}). Consequently, the supplemented cows had higher fertility after the first postpartum artificial insemination and a higher percentage of pregnancies at the first service.

It is concluded that energy-mineral supplementation, fitted to the production conditions of the Andean region of Ecuador, during the gestation-lactation transition period, positively affects the metabolic profile and reproductive indicators.

References

Aardema, H., Helena, T.A. & Peter, L.A.M. 2019. "An overview on how cumulus cells interact with the oocyte in a condition with elevated NEFA levels in dairy cows". Animal Reproduction Science, 207: 131-137, ISSN: 0378-4320, DOI: https://doi.org/10.1016/j.anireprosci.2019.06.003.

Arthington, J.D. & Brown, W.F. 2005. "Estimation of feeding value of four tropical forage species at two stages of maturity". Journal of Animal Science, 83(7): 1726-1731, ISSN: 1525-3163, DOI: https://doi.org/10.2527/2005.8371726x.

Balarezo, L.R., García-Díaz, J.R., Hernández-Barreto, M.A. & García-López, R. 2016. "Metabolic and reproductive state of Holstein cattle in the Carchi region, Ecuador". Cuban Journal of Agricultural Science, 50(3): 381-392, ISSN: 2079-3480.

Balarezo, L.R, García-Díaz, J.R., Hernández-Barreto, M.A. &Vargas-Hernández, S. 2018. "Uterine Involution in Hosltein cows in the province of Carchi, Ecuador”. Revista MVZ Córdoba, 23(2): 6649-6659, ISSN: 0122-0268, DOI: https://doi.org/10.21897/rmvz.1339.

Brito, R., Blanco, G.S., Calderón, R., Preval, B. & Campo, E. 2010. Patología de la Reproducción Animal. 2^nd Ed. Ed. Félix Varela. La Habana, Cuba, pp. 1-41, ISBN: 978-959-07-1279-1.

Carvalho, P.D., Souza, H.A., Amundson, C.M., Hackbart, M.J., Fuenzalida, J.M., Ayres, H.M., Dresch, R.A., Vieira, M.L., Guenther, J.N., Grummer, R.R., Fricke, M.P., Shaver, D.R. & Wiltbank, C.M. 2014. "Relationships between fertility and postpartum changes in body condition and body weight in lactating dairy cows". Journal of Dairy Science, 97(6): 1-18, ISSN: 1525-3198, DOI: https://doi.org/10.3168/jds.2013-7809.

Castro, N., Kawashima, C., Van Dorland, H.A., Morel, I., Miyamoto, A. & Bruckmaier, R.M. 2012. "Metabolic and energy status during the dry period is crucial for the resumption of ovarian activity postpartum in dairy cows". Journal of Dairy Science, 95(10): 5804-5812, ISSN: 1525-3198 , DOI: https://doi.org/10.3168/jds.2012-5666.

David, D.M., Campos, R., Apráez, J.E., Velez, M. & Zambrano, G.L. 2018. "Metabolic and reproductive response to energy supplementation in dairy cows in tropical highlands". Revista de Ciencias Agrícolas, 35(2): 51-63, ISSN: 2256-2273, DOI: https://dx.doi.org/10.22267/rcia.183502.91.

Djokovic, R.D., Kurcubic, V.S. & Ilic, Z.Z. 2014. "Blood serum levels of macro-and micronutrients in transition and full lactations cows". Bulgarian Journal of Agricultural Science, 20(3): 715-720, ISSN: 1310-0351.

Esposito, G., Irons, C.P. & Webb, C.E. 2014. "Interactions between negative energy balance, metabolic diseases, uterine health and immune response in transition dairy cows". Animal Reproduction Science, 144(3-4): 60-71, ISSN: 0378-4320, DOI: https://doi.org/10.1016/j.anireprosci.2013.11.007.

Evans, J.R. & Henshaw, K. 2008. "Antioxidant vitamin and mineral supplements for preventing age-related macular degeneration. Review". Cochrane Database of Systematic Reviews, (1): CD000253, ISSN: 1469-493X, DOI: https://doi.org/10.1002/14651858.CD000253.pub2.

García, J.R., Cuesta, M., García-López, R., Quiñones, R., Figueredo, J.M., Faure, R., Pedroso, R. & Mollineda, A. 2010. "Characterization of the content of microelements in the soil-plant-animal system and its influence on cattle reproduction in the central region of Cuba". Cuban Journal of Agricultural Science, 44(3): 227-231, ISSN: 2079-3480.

García, J.R., Cuesta, M., Pedroso, R., Rodríguez, J., Gutiérrez, M., Mollineda, A., Figueredo, J.M. & Quiñones, R. 2007. "Suplementación parenteral de cobre en vacas gestantes: Efecto sobre postparto y terneros". Revista MVZ Córdoba, 12(2): 985-995, ISSN: 0122-0268, DOI: https://doi.org/10.21897/rmvz.419.

García, J.R., Cuesta, M., Silveira, E., Quiñones, R., Hernández, M. & Mollineda, A. 2011. "Desequilibrios metabólicos con especial referencia a las carenciales de minerales asociadas a problemas reproductivos en vacas lecheras de Cuba". Revista Electrónica de Veterinaria REDVET, 12(12): 1-12, ISSN: 1695-7504.

García, J.R., Munyori, H., Cuesta, M., Quiñones, R., Figueredo, J.M., Noval, E. & Mollineda, A. 2012. "Therapeutic efficacy and pharmacological safety of parenteral supplementation of different concentrations of copper in cows". Archiv Tierzucht, 55(1): 25-35, ISSN: 2363-9822, DOI: https://doi.org/10.5194/aab-55-25-2012.

García, JR., Noval-Artiles, E., Pérez-Bello, A., Hernández-Barreto, M. & Pérez-González, Y. 2017. "Effects of copper parenteral supplementation on the weight gain in fattening bulls". Revista MVZ Córdoba, 22(2): 5820-5827, ISSN: 0122-0268, DOI: http://dx.doi.org/10.21897/rmvz.1009.

Gebhard, S., Ronimus, R.S. & Morgan, H.W. 2001. "Inhibition of phosphofructokinases by copper (II) ". FEMS Microbiology Letters, 197(1): 105-109, ISSN: 1574-6968, DOI: https://doi.org/10.1111/j.1574-6968.2001.tb10590.x.

Griffiths, L.M., Loeffler, S.H., Socha, M.T., Tomlinson, D.J. & Johnson, A.B. 2007. "Effects of supplementing complexed zinc, manganese, copper and cobalt on lactation and reproductive performance of intensively grazed lactating dairy cattle on the South Island of New Zealand". Animal Feed Science and Technology, 137(1-2): 69-83, ISSN: 0377-8401, DOI: https://doi.org/10.1016/j.anifeedsci.2006.10.006.

LeBlanc, S. 2010. "Monitoring metabolic health of dairy cattle in the transition period". Journal of Reproduction and Development, 56(Suppl): S29-35, ISSN: 1348-4400, DOI: https://doi.org/10.1262/jrd.1056s29.

Macrae, A. 2019. "Assessment of energy balance in dairy cattle". Livestock, 24(5): 229-235, ISSN: 2053-0870, DOI: https://doi.org/10.12968/live.2019.24.5.229.

McDowell, L.R. & Arthington. 2005. Minerales para rumiantes en pastoreo en regiones tropicales. 4^th Ed. Ed. Universidad de La Florida. Departamento de Zootecnia, Universidad de la Florida. Gainesville, Florida, U.S.A., p. 91.

Moyano, M.E., Rodríguez, C.E. 2014. "Suplementación energética y su efecto en el nivel de colesterol y el perfil hormonal preovulatorio en vacas". Revista de Salud Animal, 36(2): 90-96, ISSN: 0253-570X.

Noval, E., García-Díaz, J.R., García-López, R. & Jiménez-Pérez, J. 2016. "Efecto de diferentes dosis de un compuesto inyectable de Cu, Zn y Mn en parámetros bioproductivos de la vaca lechera". Cuban Journal of Agricultural Science, 50(2): 371-380, ISSN: 2079-3480.

NRC (National Research Council). 2001. Nutrient Requirements of Dairy Cattle. 7^th Rev. Ed. Ed. The National Academies Press. Washington D.C., U.S.A, p. 381, ISBN: 978-0-309-06997-7, DOI: https://doi.org/10.17226/9825.

Omur, A., Kirbas, A., Aksu, E., Kandemir, F., Dorman, E., Kaynar, O. & Ucar, O. 2016. "Effects of antioxidant vitamins (A, D, E) and trace elements (Cu, Mn, Se, Zn) on some metabolic and reproductive profiles in dairy cows during transition period". Polish Journal of Veterinary Science, 19(4): 697-706, ISSN: 1505-1773, DOI: https://doi.org/10.1515/pjvs-2016-0088.

Quintela, L.A., Díaz, C., Becerra, J.J., Alonso, G., Gracia, S. & Herradón, P.G. 2008. "Papel del β-caroteno y la vitamina A en la reproducción en el ganado vacuno: revisión". ITEA, Información Técnica Económica Agraria, 104(3): 399-410, ISSN: 1699-6887.

Rodenburg, J. 2004. Body Condition Scoring of Dairy Cattle. OMAFRA Factsheet. [online] Order (92-122): 414-420. [Consulted 2010September 4].

Shonewille, J.T. 2013. "Magnesium in dairy cow nutrition: an overview". Plant and Soil, 368: 167-178, ISSN: 1573-5036, DOI: https://doi.org/10.1007/s11104-013-1665-5.

StatPoint Technologies. 2010. Statgraphics Centurion User Manual. Version 16.1 [Windows]. Statgraphics Technologies, Inc. The Plains, Virginia, U.S.A, Available: < https://www.statgraphics.com/download-statgraphics-centurion-xvi>.

Walsh, R.B., Kelton, D.F., Duffield, T.F., Leslie, K.E., Walton, J.S. & LeBlanc, S.J. 2007. "Prevalence and risk factors for postpartum anovulatory condition in dairy cows". Journal of Dairy Science, 90(1): 315-324, ISSN: 1525-3198, DOI: https://doi.org/10.3168/jds.S0022-0302(07)72632-2.

Received: January 15, 2021; Accepted: February 17, 2021

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Conflict of interest: The authors declare that there are no conflicts of interests among them

Author´s contribution: Luis Rodrigo Balarezo-Urresta: Original idea, design and conducting the experiment, data analysis, manuscript writing. Ernesto Noval-Artiles: Conducting the experiment, data analysis, manuscript writing. Juan Ramón García Díaz: Conducting the experiment, data analysis, manuscript writing.