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# Evaporative water loss from dairy cows in climate-controlled respiration chambers

Open AccessPublished:January 09, 2023

## ABSTRACT

The effects of ambient temperature (AT) on total evaporative water loss from dairy cows at different relative humidity (RH) and air velocity (AV) levels were studied. Twenty Holstein dairy cows with an average parity of 2.0 ± 0.7 and body weight of 687 ± 46 kg participated in the study. Two climate-controlled respiration chambers were used. The experimental indoor climate was programmed to follow a diurnal pattern with AT at night being 9°C lower than during the day. Night AT was gradually increased from 7 to 21°C and day AT was increased from 16°C to 30°C within an 8-d period, both with an incremental change of 2°C/d. The effect of 3 RH levels with a diurnal pattern were studied as well, with low values during the day and high values during the night: low (day, 30%; night, 50%), medium (day, 45%; night, 70%), and high (day, 60%; night, 90%). The effects of AV were studied during the daytime at 3 levels: no fan (0.1 m/s), fan at medium speed (1.0 m/s), and fan at high speed (1.5 m/s). The medium and high AV levels were only combined with medium RH. In total, there were 5 treatments with 4 replicates each. The animals had free access to feed and water. Based on the water balance principle inside the respiration chambers, the total evaporative water loss from dairy cows at a daily level was quantified by measuring the mass of water in the incoming and outgoing air, condensed water, added water from a humidifier, and evaporative water from a wet floor, drinking bowl, manure reservoir, and water bucket. Water evaporation from a sample skin area was measured with a ventilated skin box, and water evaporation, through respiration with a face mask. The results show that RH/AV levels had no significant effect on total evaporative water loss, whereas the interaction effect between RH/AV with AT was significant. Cows at a high RH had a tendency for a lower increasing rate of evaporative water loss compared with cows at a low RH (0.61 vs. 0.79 kg/d per 1°C increase of AT). Cows at medium and high AV levels had a greater increasing rate than cows at low AV (0.91 and 0.95 vs. 0.71 kg/d per 1°C increase of AT, respectively). The increase of evaporative heat loss from dairy cows was mainly a result of the increase in evaporation (of sweat) from the skin. The skin water evaporation determined with the water balance method (less evaporation from respiration) and the ventilated skin box method showed no significant difference. The implication of this study is that cows at a high AT depend mainly on evaporative cooling from the skin. The ventilated skin box method, measuring only a small part of the skin during a short period during the day, can be a convenient and accurate way to determine the total cutaneous evaporative water loss from cows.

## INTRODUCTION

Evaporative heat loss is the main thermoregulatory mechanism of cows under high air temperature conditions because sensible heat loss is limited by the small temperature gradient between body surface and warm environment (
• Taneja G.C.
Sweating in cattle: I. Cutaneous evaporative losses in calves and its relationship with respiratory evaporative loss and skin and rectal temperatures.
;
• Gebremedhin K.G.
• Wu B.
A model of evaporative cooling of wet skin surface and fur layer.
). Thus, a cow's ability to endure hot environments is dependent on the amount of heat it can dissipate via evaporation, either by sweat from the skin (
• Gebremedhin K.G.
• Hillman P.E.
• Lee C.N.
• Collier R.J.
• Willard S.T.
• Arthington J.D.
• Brown-Brandl T.M.
Sweating rates of dairy cows and beef heifers in hot conditions.
) or through the respiratory system (
• da Silva R.G.
• Maia A.S.C.
• de Macedo Costa L.L.
• de Queiroz J.P.A.F.
Latent heat loss of dairy cows in an equatorial semi-arid environment.
). The most current reliable technique to determine the total evaporative water loss is to observe the BW change within a certain duration in response to a thermoregulatory sweating/respiration stimulus (
• Finch V.A.
• Bennett I.
• Holmes C.
Sweating response in cattle and its relation to rectal temperature, tolerance of sun and metabolic rate.
;
• Holmes C.R.
Application of a weighing system for measuring total evaporative water loses in large ruminants.
;
• Cheuvront S.N.
• Kenefick R.W.
CORP: Improving the status quo for measuring whole body sweat losses.
;
• Castro P.A.
• Campos Maia A.S.
• de França Carvalho Fonsêca V.
• Carol de Melo Costa C.
• Nascimento S.T.
• Simão B.R.
• Ruggieri A.C.
• Gomes da Silva R.
Comparative methods analysis on rates of cutaneous evaporative water loss (CEWL) in cattle.
), which requires costly high-precision weighing scales and careful work. Therefore, most available data in recent literature on moisture evaporated from skin in dairy cows are primarily from studies conducting measurements on a small area of the skin surface, using paper disks, ventilated capsules, or handheld electronic calorimeters (
• Berman A.
Influence of some factors on the relative evaporation rate from the skin of cattle.
;
• Hillman P.
• Gebremedhin K.
• Parkhurst A.
• Fuquay J.
• Willard S.
Evaporative and convective cooling of cows in a hot and humid environment.
;
• Maia A.S.C.
• daSilva R.G.
• Battiston Loureiro C.M.
Sensible and latent heat loss from the body surface of Holstein cows in a tropical environment.
;
• Gebremedhin K.G.
• Hillman P.E.
• Lee C.N.
• Collier R.J.
• Willard S.T.
• Arthington J.D.
• Brown-Brandl T.M.
Sweating rates of dairy cows and beef heifers in hot conditions.
;
• de Souza Jr., J.B.F.
• de Queiroz J.P.A.F.
• dos Santos V.J.S.
• Dantas M.R.T.
• de Lima R.N.
• de Oliveira Lima P.
• de Macedo Costa L.L.
Cutaneous evaporative thermolysis and hair coat surface temperature of calves evaluated with the aid of a gas analyzer and infrared thermography.
).
However, local measurements of the sweating rate can have limitations as a result of a small sample area, a short measurement time, and measurements taken on a shaved skin or under still air conditions.
• Berman A.
Influence of some factors on the relative evaporation rate from the skin of cattle.
and
• Gebremedhin K.G.
• Lee C.N.
• Hillman P.E.
• Collier R.J.
Physiological responses of dairy cows during extended solar exposure.
found there was a significant difference in sweating rate between black and white hair coats, and
• Gebremedhin K.G.
• Lee C.N.
• Hillman P.E.
• Collier R.J.
Physiological responses of dairy cows during extended solar exposure.
noted that cows sweat in a cyclic pattern. It is well known that the entire body surface of an animal is involved in thermal exchanges with the environment (
• Turnpenny J.R.
• McArthur A.J.
• Clark J.A.
• Wathes C.M.
Thermal balance of livestock: 1. A parsimonious model.
). In this case, estimating total cutaneous evaporative water loss using the local sweating rate might have a large inaccuracy, which would be even worse when multiplying the body surface area estimated from empirical equations (
• Berman A.
Effects of body surface area estimates on predicted energy requirements and heat stress.
). Although measuring the evaporative water loss from the whole body surface of dairy cows under different conditions is a challenge, it is of great interest and can be helpful for validating the previously mentioned measurement methods and for determining a better estimation of the contribution of the latent heat loss from the skin to total heat loss, which is important information when applying cooling systems.
The objectives of this study were (1) to determine how dairy cows adjust their total evaporative water loss at different temperatures, relative humidity (RH), and air velocity (AV); and (2) to investigate whether the local cutaneous evaporative water loss measured using a ventilated skin box was in agreement with the total cutaneous evaporative water loss derived using the water balance method. We hypothesized that total evaporative water loss at an increasing ambient temperature (AT) is affected by different RH and AV levels, and cutaneous evaporative water loss measured with the ventilated skin box on a small area of the skin surface might reveal large discrepancies when compared with the total cutaneous evaporative water loss derived from the water balance method.

## MATERIALS AND METHODS

### Experimental Design

The experiment was conducted in 2021, approved by the Institutional Animal Care and Use Committee of Wageningen University & Research (Wageningen, the Netherlands), and in accordance with Dutch law (project no. 2019.D-0032). Twenty Holstein-Friesian dairy cows were used with an average milk yield (±SD) of 30.0 ± 4.7 kg/d, 206 ± 39 DIM, 687 ± 46 kg BW, and parity of 2.0 ± 0.7. Nineteen cows were pregnant at an average of 105 ± 38 d. There were 4 cows in each treatment, and cows were grouped in 4 blocks of 5 cows based on parity and expected milk yield. Each cow within a block was assigned randomly to 1 of the 5 treatments. We decided to use 4 animals in each treatment because we are aware of ethical considerations to minimize the number of cows in climate-controlled respiration chambers (CRC) (
• Lakens D.
Sample size justification.
). This sample size is common in dairy cow studies done in respiration chambers (
• Perano K.M.
• Usack J.G.
• Angenent L.T.
• Gebremedhin K.G.
Production and physiological responses of heat-stressed lactating dairy cattle to conductive cooling.
;
• Hou Y.
• Zhang L.
• Dong R.Y.
• Liang M.Y.
• Lu Y.
• Sun X.Q.
• Zhao X.
Comparing responses of dairy cows to short-term and long-term heat stress in climate-controlled chambers.
), and this study focused mainly on the underlying evaporative water loss processes and their time dynamics, not on comparing the specific treatment effects.
Cows were subjected to a 3-d adaptation period and a subsequent 8-d experimental period in the CRC with a specific treatment consisting of combinations of AT, RH, and AV. The AT inside the CRC was increased gradually from 7 to 21°C at night and from 16 to 30°C during the day within 8 d (by steps of 2°C per day for both nighttime and daytime temperatures), as shown in Figure 1. Three levels of RH during the day and night were RH_l (low), 30% during the day and 50% during the night; RH_m (medium), 45% during the day and 70% during the night; and RH_h (high), 60% during the day and 90% during the night. At nighttime, AV was kept at natural speed (AV_l; 0.1 m/s). In the daytime, 3 AV levels were applied: AV_l (0.1 m/s), AV_m (1.0 m/s), or AV_h (1.5 m/s). For AV_m and AV_h, the AT was started 2°C higher (from 18 to 32°C) than AV_l. AV_m and AV_h were only combined with RH_m, resulting in 5 treatments. The AT, RH, and AV conditions for a 3-d adaptation period in the CRC were set and controlled the same as the first day of the corresponding experimental period. A more detailed description can be found in a previous study (
• Zhou M.
• Aarnink A.J.A.
• Huynh T.T.T.
• van Dixhoorn I.D.E.
• Groot Koerkamp P.W.G.
Effects of increasing air temperature on physiological and productive responses of dairy cows at different relative humidity and air velocity levels.
). Twenty cows were grouped in 4 blocks of 5 cows based on parity and expected milk yield. Each cow within a block was assigned randomly to 1 of the 5 treatments. The cows were tied up loosely, which means they could move forward/backward and lie down easily. The flooring material was rubber mats at the front side and rubber-covered metal grilles at the back side. The cows received feed ad libitum via a feed trough fixed in front of the cubicle, and water via a drinking bowl at the left side of the head of the cow. The diet was formulated to meet or exceed the nutritional requirements of lactating Holstein cows according to the Dutch system (
• CVB
CVB Table Ruminants 2008. Series 43.
).

### Climate-Controlled Respiration Chamber

In this study, 2 identical CRC were used. Each chamber was split into 2 individual airtight compartments with thin walls equipped with a transparent windows to allow audio and visual contact between 2 cows and thereby reduce the effects of social isolation on their behavior. Each compartment had a volume of 34.5 m3 and a dimension of length × width × height (4.5 × 2.7 × 2.8 m) as described in detail by
• Gerrits W.
• Labussière E.
Indirect Calorimetry: Techniques, Computations and Applications.
. In each compartment, RH was monitored by 1 relative humidity sensor (accuracy, ≤ ±2%; resolution, = 0.0001%; Novasina Hygrodat100, Novasina AG), and AT was monitored by 5 PT100 temperature sensors (accuracy, ≤ ±0.1°C; resolution, 0.0005°C; Sensor Data BV) distributed evenly around the room at animal height as described in detail in
• Zhou M.
• Aarnink A.J.A.
• Huynh T.T.T.
• van Dixhoorn I.D.E.
• Groot Koerkamp P.W.G.
Effects of increasing air temperature on physiological and productive responses of dairy cows at different relative humidity and air velocity levels.
. The different RH levels were achieved by means of a humidifier (ENS-4800-P, Stulz) or a dehumidifier (koeltechniek, Nijssen), and the circulating air was heated or cooled depending on the deviation from set point temperatures, the control mechanism of which is reported by
• Gerrits W.
• Labussière E.
Indirect Calorimetry: Techniques, Computations and Applications.
. The settings for AV were achieved using a ventilator (diameter, 500 mm; Professional Fans, model 8879, HBM Machines BV) that was fixed on the ceiling of the chamber, 2.5 m above the floor (Figure 2), so that the airflow moved around the axial body length of the cow from back to front. The chambers were artificially lit for 16 h daylight (390 to 440 lx, 0500 to 2100 h) and 8 h nightlight (35 to 40 lx, 2100 to 0500 h).

### Data Collection

#### CRC Condition.

Throughout the 8-d experimental period, the temperature and RH of the chamber compartments were recorded continuously at 30-s intervals automatically. Using a handheld anemometer (Testo 5-412-983, Testo SE & Co. KGaA), actual AV was measured manually 3 times a day at 5 locations with about a 5-cm distance around the cow's body surface (neck, middle trunk, rump, and both lateral sides) for 30 s each.

#### Metabolic Heat Production.

Throughout the 8-d experimental period, CO2 and CH4 production, and O2 consumption were measured at 12-min intervals and recorded automatically, as described by
• Gerrits W.
• Labussière E.
Indirect Calorimetry: Techniques, Computations and Applications.
. Based on these data, metabolic heat production was calculated using the equation according to
• McLean J.A.
On the calculation of heat production from open-circuit calorimetric measurements.
.

#### Evaporative Water Loss.

To quantify the total evaporative water loss from the cow during the experimental period, a complete water balance inside the CRC (Figure 2) was calculated as follows for each cow:
evaporative water loss = (A + B) – (C + D + E + F + G + H),

where evaporative water loss is the total evaporative water loss of a cow (i.e., water evaporated from the skin surface and from the respiratory track, measured in kg/d); A is the mass of water in the outgoing air, calculated daily by measuring the volume and humidity of the outgoing air every 30 s; and B is the amount of water that condensed in the heat exchangers. Water was collected in a tank outside the chamber, which was weighed and recorded daily. C is the mass of water in the incoming air, calculated daily by measuring the volume and humidity of the incoming air every 30 s; D is the amount of water evaporated from a bucket with a known surface area (0.16 m2), which was measured and recorded continuously at 5-s intervals. The evaporated water was then divided by the surface area of the bucket to calculate the evaporation rate (kg × m−2) for different periods, which was then used to calculate E, F, and G. E is the volume of water evaporated from wet, solid floor. The wet area on the solid floor was determined 3 times a day (at the same measurement times of local cutaneous evaporative water loss) by estimating the proportion of the solid floor wetted with drinking water and urine during different periods using the method of
• Huynh T.T.T.
• Aarnink A.J.A.
• Heetkamp M.J.W.
• Verstegen M.W.A.
• Kemp B.
Evaporative heat loss from group-housed growing pigs at high ambient temperatures.
. F and G are the amount of water evaporated from the manure reservoir and drinking water bowl, respectively. E, F, and G were calculated by multiplying the surface area of the wet floor, reservoir, and bowl with the water evaporation rate per square meter (D/0.16). H is the volume of water that was added to maintain the setup humidity in the CRC. The volume of water sprayed into the air by a humidifier was measured daily. To calculate the energy used for evaporation, the enthalpy of vaporization at 35°C was used: 2,417.9 kJ heat/1 kg water.
Three times daily (0600, 1000, and 1800 h), local cutaneous evaporative water loss from a sampling area at the belly was measured using a ventilated skin box, and evaporative water loss through respiration was measured using a face mask and a nose cup. The ventilated skin box was designed with a sampling box with 2 temperature and RH sensors mounted on both the inlet and outlet of the box, and an air suckling pump was connected to the outlet, similar to the one described by
• Gebremedhin K.G.
• Hillman P.E.
• Lee C.N.
• Collier R.J.
• Willard S.T.
• Arthington J.D.
• Brown-Brandl T.M.
Sweating rates of dairy cows and beef heifers in hot conditions.
. The net heat loss or gain from the sampling area was calculated from the property differences of the incoming and outgoing air. Similarly, the respiratory heat loss was calculated from the property differences of the inhaled and exhaled air. A more detailed description can be found in
• Zhou M.
• Huynh T.T.T.
• Groot Koerkamp P.W.G.
• van Dixhoorn I.D.E.
• Amon T.
• Aarnink A.J.A.
Effects of increasing air temperature on skin and respiration heat loss from dairy cows at different relative humidity and air velocity levels.
. To determine the local cutaneous evaporative water loss measured by the ventilated skin box and the respiratory water loss at a daily level, the local cutaneous evaporative water loss and respiratory water loss during the day condition (1000 to 1900 h) was calculated using the mean of the values measured at 1000 and 1800 h. During the increasing temperature condition (0700 to 1000 h), they were calculated using the mean of the values measured at 0600 and 1000 h. During the decreasing temperature condition (1900 to 2200 h), they were calculated using the mean of 2 values measured at 1800 and 2200 h (previous day). During the night condition (2200 to 0700 h), they were calculated using the value measured at 0600 h, as illustrated in Figure 3. The total cutaneous evaporative water loss (kg/d) was calculated by subtracting the daily respiratory water loss from the total evaporative water loss (calculated using the water balance method), and this was converted to grams per square meter per hour by applying the body surface area (0.14BW0.57) as estimated according to
• Brody S.
Bioenergetics and Growth: With Special Reference to the Efficiency Complex in Domestic Animals.
.

### Statistical Analysis

One cow (receiving treatment, RH_m and AV_m) was excluded from the experiment because of mastitis. Descriptive statistics are presented in Table 1 for different elements used for calculating the water balance under different treatments. Statistical procedures were carried out using SAS 9.4 (SAS Institute Inc.). All data were screened to confirm the normality and homoskedasticity of variances. The MIXED procedure was used to investigate the fixed effects of AT, treatments (combinations of RH and AV), and their interaction. The baseline milk yield (average milk yield from first 2 d) was included as covariate; cow was included as a random effect.
Table 1The mean (±SE) of different elements for calculating the total evaporative water loss (EWL, kg/d) from dairy cows under different treatments
Treatment
Treatments during the day and night: RH_l, 30% to 50%; RH_m, 45% to 70%; RH_h, 60% to 90%; AV_l, 0.1 to 0.1 m/s; AV_m, 1.0 to 0.1 m/s; AV_h, 1.5 to 0.1 m/s. RH = relative humidity; l = low; AV = air velocity; m = medium; h = high.
Air water out/in
The difference between the amount of water in the outgoing air and the incoming air (kg/d).
Condensed water
The amount of water condensed in the heat exchangers (kg/d).
Humidifier water
The amount of water sprayed into the air by a humidifier to maintain the setup humidity (kg/d).
EWL from wet floor
The amount of water evaporated from wet, solid floor (kg/d).
EWL elsewhere
The amount of water evaporated from a drinking bowl, manure reservoir, and water bucket (kg/d).
EWL from cows
The amount of total evaporative water loss from the skin surface and respiration from dairy cows (kg/d).
RH_l × AV_l5.03 ± 0.2120.8 ± 0.9700.622 ± 0.0743.68 ± 0.2321.5 ± 0.82
RH_m × AV_l7.15 ± 0.5820.1 ± 0.5200.960 ± 0.0773.15 ± 0.1423.1 ± 0.72
RH_h × AV_l10.1 ± 0.8213.7 ± 0.642.78 ± 0.4410.284 ± 0.0592.21 ± 0.1318.5 ± 0.67
RH_m × AV_m7.36 ± 0.5622.2 ± 0.9401.10 ± 0.214.72 ± 0.2023.7 ± 1.1
RH_m × AV_h7.90 ± 0.5321.9 ± 0.6500.731 ± 0.134.56 ± 0.1824.5 ± 0.84
1 Treatments during the day and night: RH_l, 30% to 50%; RH_m, 45% to 70%; RH_h, 60% to 90%; AV_l, 0.1 to 0.1 m/s; AV_m, 1.0 to 0.1 m/s; AV_h, 1.5 to 0.1 m/s. RH = relative humidity; l = low; AV = air velocity; m = medium; h = high.
2 The difference between the amount of water in the outgoing air and the incoming air (kg/d).
3 The amount of water condensed in the heat exchangers (kg/d).
4 The amount of water sprayed into the air by a humidifier to maintain the setup humidity (kg/d).
5 The amount of water evaporated from wet, solid floor (kg/d).
6 The amount of water evaporated from a drinking bowl, manure reservoir, and water bucket (kg/d).
7 The amount of total evaporative water loss from the skin surface and respiration from dairy cows (kg/d).
Cutaneous evaporative water loss rates were analyzed using the least squares method by fitting generalized linear models (PROC GLM), including the fixed effects of 2 methods (derived by the local measurement using the ventilated skin box and the water balance method), AT, and the interaction effect between the 2 methods and AT. The root mean square deviation (RMSD), the mean bias (MB), and the coefficient of determination (R2) were used to determine the deviation between measured evaporative water loss by the ventilated skin box and by the water balance method. The differences between the 2 methods were quantified by RMSD as follows:
$RMSD=∑(xi−yi)2n,$

where xi and yi are cutaneous evaporative water loss rates derived by the ventilated skin box and water balance method, respectively; and n is the number of measurements.
MB is the average bias and calculated as
$MB=∑(xi−yi)n.$

R2 is calculated as
$R2=1−∑(xi−yi)2∑(xi−x¯)2,$

where $x¯$ is the mean of xi (i = 1, 2 , … , n).

## RESULTS

### Effect of AT, RH, and AV

Total evaporative water loss from cows (respiration and sweating) generally increased with increasing AT for individual cows, as shown in Figure 4, and the difference between individual cows was substantial. Milk yield tended to be associated positively with evaporative water loss (P = 0.052). RH/AV had no significant effect on evaporative water loss (P = 0.26), whereas the interaction effect between RH/AV with AT was significant (P = 0.0024). Specifically, at a low AV level, cows at a high RH had a tendency for a lower increasing rate (0.61 kg/d per 1°C increase of AT) of evaporative water loss compared with cows at a low RH (0.79 kg/d per 1°C increase of AT, P = 0.065). At a medium RH, cows at medium and high AV levels had a greater increasing rate (0.91 and 0.95 kg/d per 1°C increase of AT, respectively) than cows at a low AV (0.71 kg/d per 1°C increase of AT, P < 0.05). Figure 5 shows the partitioning of metabolic heat production in which the total evaporative heat loss increased and the sensible heat loss (the difference between total heat production and latent heat loss, assuming the cows were in heat balance) decreased with increasing AT. Figure 5 also shows that the increase of evaporative heat loss from dairy cows was mainly a result of the increase of evaporation (of sweat) from the skin. The respiratory evaporative heat loss measured with a face mask increased only slightly with increasing AT.

### Comparison of Cutaneous Evaporative Water Loss Rate Measured Using 2 Methods

There was no significant difference in the rate of cutaneous evaporative water loss between the 2 methods used (P = 0.387). The RMSD and MB of the rates of cutaneous evaporative water loss obtained by water balance method and ventilated skin box were 39.1 and 0.44 g × m−2 × h−1, equaling 42% and 0.5% of the total cutaneous water loss obtained by the first method, respectively. The values measured using the ventilated skin box were slightly greater than those calculated using the water balance method. R2 was equal to 0.87 (Figure 6) using the correlation-regression approach (intercept = 0), which means 87% of the variance of the rates of cutaneous evaporative water loss calculated using the water balance method can be explained by the values measured using the ventilated skin box.

## DISCUSSION

Quantification of the total evaporative water loss is of interest to study the different routes of heat loss in dairy cows. This information also gives insight into the effect of different cooling systems on additional heat losses. To the best of our knowledge, this is the first study to determine the total evaporative water loss rate from dairy cows at a daily level, which can avoid some sources of errors associated with different sweating rates between different skin regions, and cyclic sweating patterns (
• Berman A.
Influence of some factors on the relative evaporation rate from the skin of cattle.
;
• Gebremedhin K.G.
• Hillman P.E.
• Lee C.N.
• Collier R.J.
• Willard S.T.
• Arthington J.D.
• Brown-Brandl T.M.
Sweating rates of dairy cows and beef heifers in hot conditions.
;
• Liang B.
• Parkhurst A.
• Gebremedhin K.
• Lee C.
• Collier R.
• Hillman P.
Using time series to study dynamics of sweat rates of Holstein cows exposed to initial and prolonged solar heat stress.
;
• de Souza Jr., J.B.F.
• de Queiroz J.P.A.F.
• dos Santos V.J.S.
• Dantas M.R.T.
• de Lima R.N.
• de Oliveira Lima P.
• de Macedo Costa L.L.
Cutaneous evaporative thermolysis and hair coat surface temperature of calves evaluated with the aid of a gas analyzer and infrared thermography.
). With the design of our experiment, we were able to estimate the total evaporative water loss from cows as well as separate it between skin evaporation and respiratory evaporation.

### Effect of AT, RH, and AV

The total evaporative water loss rate increased as the environmental temperature rose. This is a consequence of the decreased temperature gradient between the skin surface and surrounding air, causing a decrease in sensible heat loss that needs to be compensated by an increase in latent heat loss. We included baseline milk yield as a covariate when analyzing the total evaporative water loss because cows producing more milk normally have greater metabolic heat production (
• Ravagnolo O.
• Misztal I.
• Hoogenboom G.
Genetic component of heat stress in dairy cattle, development of heat index function.
) and thus a need for greater total heat loss.
• Castro P.A.
• Campos Maia A.S.
• de França Carvalho Fonsêca V.
• Carol de Melo Costa C.
• Nascimento S.T.
• Simão B.R.
• Ruggieri A.C.
• Gomes da Silva R.
Comparative methods analysis on rates of cutaneous evaporative water loss (CEWL) in cattle.
used a weighing system as a gold standard method to quantify the total evaporative water loss by observing acute changes in body mass. This method, however, has limitations because it neglects gas losses from the body (
• Cheuvront S.N.
• Kenefick R.W.
CORP: Improving the status quo for measuring whole body sweat losses.
), which is a larger error source in cows because of rumination activity compared with human beings. The range of the cutaneous evaporative water loss rate measured using this weighing system over a period of 2 h was 54 to 375 g × m−2 × h−1 within AT around 26 to 34°C (
• Castro P.A.
• Campos Maia A.S.
• de França Carvalho Fonsêca V.
• Carol de Melo Costa C.
• Nascimento S.T.
• Simão B.R.
• Ruggieri A.C.
• Gomes da Silva R.
Comparative methods analysis on rates of cutaneous evaporative water loss (CEWL) in cattle.
). In our study, the range was between 12.8 and 183.4 g × m−2 × h−1, which was the daily mean of the cutaneous water loss under higher AT during the day (16 to 32°C) and lower AT during the night (7 to 23°C). The mass loss by gas exchange (the difference between the production of CO2 and CH4, and the consumption of O2) was calculated in this study to vary from 28.5 to 41.1 g × m−2 × h−1. The calculated gas loss weight could account for 8% to 76% of the cutaneous water loss measured by
• Castro P.A.
• Campos Maia A.S.
• de França Carvalho Fonsêca V.
• Carol de Melo Costa C.
• Nascimento S.T.
• Simão B.R.
• Ruggieri A.C.
• Gomes da Silva R.
Comparative methods analysis on rates of cutaneous evaporative water loss (CEWL) in cattle.
, causing considerable errors when this gold standard method is used as a reference value for comparing different measurement methods, especially under cool conditions.
In our study, we found that cows had more or less equal heat production and evaporative heat loss at low and medium RH levels. This could explain why the inflection point temperatures at which rectal temperature started to increase at low and medium RH levels were similar (25.3 and 25.9°C, respectively) (
• Zhou M.
• Aarnink A.J.A.
• Huynh T.T.T.
• van Dixhoorn I.D.E.
• Groot Koerkamp P.W.G.
Effects of increasing air temperature on physiological and productive responses of dairy cows at different relative humidity and air velocity levels.
). However, at a high RH level, the inflection point temperature was a lot lower (20.1°C), which was likely a result of greater metabolic heat production and lower evaporative heat loss. To be specific, at high AT and high RH, evaporating sweat to ambient air became more difficult (
• Berman A.
Predicted limits for evaporative cooling in heat stress relief of cattle in warm conditions.
) as shown in Figure 5. As a result, skin temperatures at a high RH level were greater than those at low or medium RH levels, which caused more sensible heat dissipation to the ambient air. We found cows had greater evaporative heat loss at medium and high AV levels than at a low AV level, and the inflection point temperature for rectal temperature at a medium level was less than others (
• Zhou M.
• Aarnink A.J.A.
• Huynh T.T.T.
• van Dixhoorn I.D.E.
• Groot Koerkamp P.W.G.
Effects of increasing air temperature on physiological and productive responses of dairy cows at different relative humidity and air velocity levels.
). The physiological responses—rectal temperature and evaporative heat loss—could happen in a parallel, so cows going into heat stress have a higher rectal temperature and higher evaporative heat dissipation (
• da Silva R.G.
• Maia A.S.C.
Thermal balance and thermoregulation.
).
Interestingly, the increase in evaporative heat loss from dairy cows under warm conditions was mainly a result of an increase in evaporation (of sweat) from the skin. The increase in respiration rate as a response to increasing ambient temperatures seems to occur primarily to compensate for the reduced temperature gradient between inhaled and exhaled air. Therefore, evaporative skin cooling is of utmost importance when cows are exposed to heat load, although this effect was caused in part by the setup of our experiment, in which RH was independent of increasing temperature whereas, in reality, RH often decreases with increasing AT.

### Comparison of Cutaneous Evaporative Water Loss Rate Measured Using 2 Methods

The bias of the local cutaneous evaporative water loss rate obtained using the ventilated skin box with the total rate determined using the water balance method was quite small (0.5%). The random deviation was larger (42%), but that could not be accounted for only by errors in the ventilated skin box method. In our study, we selected evaporative water loss from the side belly as being representative of the whole body, referring to the study of
• Castro P.A.
• Campos Maia A.S.
• de França Carvalho Fonsêca V.
• Carol de Melo Costa C.
• Nascimento S.T.
• Simão B.R.
• Ruggieri A.C.
• Gomes da Silva R.
Comparative methods analysis on rates of cutaneous evaporative water loss (CEWL) in cattle.
, and each measurement took 10 min: 5 min for adaptation and 5 min for calculation. The rather good overall agreement is an interesting result, because there are several error sources when using the ventilated skin box to measure cutaneous evaporative water loss, as mentioned earlier: (1) the sweating rate varies between different locations (
• de Souza Jr., J.B.F.
• de Queiroz J.P.A.F.
• dos Santos V.J.S.
• Dantas M.R.T.
• de Lima R.N.
• de Oliveira Lima P.
• de Macedo Costa L.L.
Cutaneous evaporative thermolysis and hair coat surface temperature of calves evaluated with the aid of a gas analyzer and infrared thermography.
); (2) there are cyclic sweat secretion patterns, causing variations in time (
• Gebremedhin K.G.
• Lee C.N.
• Hillman P.E.
• Collier R.J.
Physiological responses of dairy cows during extended solar exposure.
); (3) the airflow rate through the ventilated box is different from the airflow surrounding the cow's trunk (
• McLean J.A.
The partition of insensible losses of body weight and heat from cattle under various climatic conditions.
); and (4) the positions (lying/standing) and activities (eating/resting) would have certain effects on sweating rate. The results from
• Castro P.A.
• Campos Maia A.S.
• de França Carvalho Fonsêca V.
• Carol de Melo Costa C.
• Nascimento S.T.
• Simão B.R.
• Ruggieri A.C.
• Gomes da Silva R.
Comparative methods analysis on rates of cutaneous evaporative water loss (CEWL) in cattle.
showed there was a significant difference (P = 0.0398) between cutaneous evaporative water loss measured using 3 different methods (weighing system, ventilated capsule, and colorimetric paper disks). Although they mentioned that the water loss determined by the weighing system was the gold standard method (
• Finch V.A.
• Bennett I.
• Holmes C.
Sweating response in cattle and its relation to rectal temperature, tolerance of sun and metabolic rate.
), we found, as mentioned earlier, that gas exchanges, which were not included in their calculations (
• Castro P.A.
• Campos Maia A.S.
• de França Carvalho Fonsêca V.
• Carol de Melo Costa C.
• Nascimento S.T.
• Simão B.R.
• Ruggieri A.C.
• Gomes da Silva R.
Comparative methods analysis on rates of cutaneous evaporative water loss (CEWL) in cattle.
), could account for a considerable amount of the weight change of a cow. As a result, the comparison between different methods became difficult.
In our study, cutaneous evaporative water loss at a daily level measured using a ventilated skin box was calculated from the weighted means of 3 measurements at different times during the day. Whether this estimation could cause a big error for representing daily cutaneous evaporative water loss is unknown. The values measured using the ventilated skin box show good agreement with those measured using the water balance method despite the fact that the cows were evaluated at a daily level with high AT during the day and lower AT during the night. Whether this finding can be applied to other conditions requires further investigation.

## CONCLUSIONS

Total evaporative water loss increased as AT rose. The RH and AV levels had no significant effect on total evaporative water loss, whereas the interaction effect between RH/AV with AT was significant. The increase in evaporative heat loss from dairy cows was mainly a result of the increase in evaporation from the skin. A comparison of data measured using a ventilated skin box with parallel data derived using the water balance method revealed no significant differences in cutaneous evaporative water loss.

## ACKNOWLEDGMENTS

This research was financially supported by the China Scholarship Council (Beijing, China), the Sino-Dutch Dairy Development Center (Beijing, China), and the Farm Technology Group (Wageningen University and Research, Wageningen, the Netherlands). The authors gratefully acknowledge technical assistance from Marcel Heetkamp, Sven Alferink, Tamme Zandstra, and the animal caretakers of the experimental facilities of “Carus” (Wageningen University and Research). The authors have not stated any conflicts of interest.

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