Measuring water holding capacity in poultry meat

Abstract

In the current scientific literature, one can find >100 different methods to evaluate water-holding capacity in fresh and cooked meat. The main concepts are based on removing some of the water by either gravity, application of pressure (e.g., centrifugal force), and heating while measuring water exudate to predict the water holding capacity (WHC) during storage, processing, cooking, and/or distribution. More sophisticated methods include nuclear magnetic resonance (NMR) in which the relaxation of water molecules within a meat protein/gel system is measured to predict how the water (75% in lean meat) will behave during processing. Overall, the number of tests reported is also so high because there are quite big variations in test conditions (e.g., 750–30,000 g for centrifugal testing). The aim of this article (outcome of a symposium on methods for poultry meat characterization) is to help the reader navigate through the different setups and suggest standardized testing based on scientific principles. The recommended WHC test is the application of low centrifugal force (750 g so sample is not permanently deformed) to a protein gel, while the sample is placed on a screen platform to avoid reabsorbing the liquid separating during the slowing down of the centrifuge. It is also recognized that some meat samples (e.g., high in fat) might require a different g-force, so it is recommended to employ both the conditions mentioned above and the lab-specific conditions. Our overall goal should always be to increase uniformity in test procedures, which will enhance our capabilities to compare results among research groups.

INTRODUCTION

The word “measurement” is used to describe a basic concept in the study of mathematics and science. It quantifies the characteristics of an object or event, which we can compare with other objects or events. Measurement is the most used word whenever we deal with the division of a quantity (Anonymous, 2022). Overall, measurements should be accurate, precise, reproducible, and acceptable. The term accuracy refers to how close a measurement is to the true or acceptable value. The term precision refers to how close 2 measurements of the same item are to each other. A simple example can be shown by describing 4 darts hitting a target. If they are all hitting the center of the target (“bullseye”), they will be described as both accurate and precise. If they are concentrated in one corner away from the bullseye, they will be described as precise but not accurate. If they are all scattered over the board, they will be described as neither accurate nor precise. If they are scattered around the board but at an even distance from the bullseye they will be described as mathematically accurate. This is an important concept in understanding what one would like to achieve within an acceptable method. The fact that there are over 100 methods to determine water holding capacity (WHC) of meat in the literature does not make the selection easy. In the industry, the term WHC is used to describe the amount of water that can be held by the meat (whole muscle or minced). The term is mainly related to the interactions among water molecules and proteins (i.e., each individual protein structure and charge, as well as the architecture produced by a group of proteins). Overall, WHC is one of the most important traits of meat quality as lean meat contains 75% water that can lost when meat is cut (called drip loss), cooked (called cooking loss) or stored due to purge. These losses directly translate to financial and sensorial properties (e.g., reduced juiciness) when processing and delivering meat to the customer. Table 1 provides some examples of conditions used to determine the WHC of different meats. It is obvious that different pressures will affect the extraction of different layers of bound water in a meat sample (see below a discussion concerning the different layers of water). When measuring WHC in meat, one should realize that lean muscle is roughly 75% water. This water can be divided into 4 main categories (Honikel, 2009). The first layer of water adjacent to the protein molecules is considered to be the bound water as it is bound electrostatically to the muscle proteins, and accounts for only 1% of the total water in the muscle (at pH 7) or in meat (at pH 5.5). The second category is the intra-myofibrillar water which represents 80% of the water in muscle and 75% in meat. The third is extra-myofibrillar and intra-cellular water at 15% and 10%, respectively. The fourth is the extracellular which is 5 and 15%, respectively. Others such as Kinsella et al. (1989) discussed 6 types of water, while Huff-Lonergan and Lonergan (2005), simplified this and discussed 3 main layers of water (Figure 1A), consisting of bound water which they described as water molecule that are tightly bound directly to the muscle proteins, immobilized water where the water molecule movement is restricted and the amount decreases with the progression of rigor mortis (pH declines which affects the charges on the proteins; see Figure 1B and further discussion below), and free water with little or no restriction on water molecule movement.

Table 1.

Examples of different conditions used for centrifugal force applications reported in the literature.

Investigators	Meat type	Mass (gram)	Force	Temperature (°C)	Time (min)	Salt (%)
Bouton et al. (1972)	Ground mutton	3–4	150,000 g	*	30–60	*
Dagbjartsson and Solberg (1972)	Ground lobster	1.0	12,000 g	*	10	*
Hermansson and Akesson (1975)	Beef and pork added with protein (raw)	20	18,000 g	*	30	*
Jauregui et al. (1981)	Ground fish	1.5	7,710 g	2	10	0
	Ground chicken	1.5	17,300 g	2	15	0
	Ground beef	1.5	30,900 g	2	30	0
Eide et al. (1982)	Minced fish	2	1,500 g	10	5	*
Hashizawa et al. (2013)	Ground broiler	1	1,000 g	4	15	0
Bowker and Zhang (2015)	Ground broiler	10	7,000 g	4	15	2.2
Liu et al. (2022)	Ground carp	2	1,590 g	*	10	0
Yang et al. (2022)	Sarcoplasmic proteins	2	3,000 g	25	1	0.01

^⁎Value not reported.

Figure 1.

Schematic of the types of water in whole muscle: (A) based on Huff-Longeran and Lonergan (2005), and (B) effect of pH on water holding capacity from Barbut (2015).

It should be pointed out that some water is held by capillary forces, which can represent a significant force in holding the water within the structure (e.g., tall trees rely on capillary forces to bring up water to height of > 50 m). Below are some numbers to illustrate this point: a capillary of 2 mm gives rise to 15 mm of water at 20°C . When the radius is reduced to 1 and 0.1 mm, the water column height is 40 and 300 mm, respectively. In any case, retaining the water in fresh meat (e.g., deboned chicken breast fillets), and in further processed cooked meat products (e.g., chicken rolls, turkey ham) is very important for both the individual consumer and the industry. For the individual consumer, the fresh meat in a package should not show any exudate as this is seen as an inferior product. The consumer would also like to have a product that can maintain its moisture during cooking, so it is not too dry and chewy. For the industry, retaining water in the meat is essential to getting higher production yields. This is a very important point for the industry wanting to produce acceptable juicy products (e.g., during cooking of meat the water holding capacity is decreasing due to protein denaturation) while also maintaining profitability by retaining the original water and any added moisture (brine/marinade) (Barbut, 2015).

Methods for Measuring WHC of Meat

There are various methods to accurately measure the dynamics of water in biological systems. However, techniques such as determining absorption isotherms, water activity, osmotic pressure, relative vapor pressure, and differential scanning calorimetry provide good data on some parameters of water, but do not provide good quantitative information that can be used to predict the actual WHC of a given lot of meat (Warner, 2014). Predicting and/or estimating the ability of meat to hold its own moisture or added water (added during further processing by tumbling/injection; commonly ranging from 15 to 50%) is very important information for the industry to use in meat formulating programs (e.g., Least Cost Formulation; software that allows exchange of meats without affecting the outcome/quality of the product), for maintaining consumer acceptance (e.g., juiciness, color), and for forecasting processing costs.

The methods that have been most consistent and useful in proving information related to WHC in meat (raw and cooked) are discussed below, including examples of the variations that have been reported by different research groups over the years. It should be pointed out that this is the reason for the existence of so many methods appearing in the scientific literature, and therefore the difficulty in comparing results even when 2 groups used the same basic method (e.g., press method) but with different conditions (e.g., different pressure, compression time, temperature, age of the meat sample). Overall, the methods can be divided into 1) gravimetric methods, 2) application of external forces (e.g., hydraulic, centrifugal), 3) application of heat (e.g., cooking loss), and 4) nondestructive estimations (e.g., hyperspectral imaging, NMR).

Gravimetric Methods

Gravimetric methods are the simplest to perform, where weight loss is determined over time (short or long period) without the application of external forces. In this case, drip loss from samples (with specified geometry, weight, and fiber orientation) suspended inside a plastic bag, or placed in special plastic tubes are collected over a certain period. Tests are performed under atmospheric pressure, and predetermined temperature and time (hours to several days). Numerous variations to this method have been reported. The main advantages are simplicity, low cost, and no need for special laboratory skills. However, the test takes a few days (slow to get results, while meat is a perishable item). Background information on animal and processing history is important to collect as it can also influence the findings (Honikel, 1998; Kapper et al., 2008). As for excising the sample from a piece of meat (e.g., chicken breast fillet), Honikel (1998) recommended having the muscle fibers oriented parallel to the long axis. However, some studies indicated the opposite, and some have not included this information in their description. An example of using the test is from Honikel et al. (1986) who studied the effect of sarcomere length on WHC in beef and pork muscles, after 7 days of storage at 0°C. Overall, they demonstrated a linear relationship at the range of 0.7 to 1.9 µm, where drip loss was 9 mL for the shorter sarcomere length, and 2 mL for the longer one, as shorter sarcomeres result in less spaces for water when the muscle is super contracted.

Application of an External Force

Various forces such as centrifugal force, pressure (positive, negative/vacuum), and capillary absorption have been applied to samples to speed up the process of proportional water release to allow prediction of WHC under regular storage conditions (e.g., refrigeration), or processing (e.g., cutting, freezing). In these cases, the time for the test is reduced from a few days to a few hours/min compared to gravimetric methods.

Centrifugal Force

A certain g-force is applied to a pre-determined amount of sample with a specific configuration. The sample is placed in a centrifuge tube, spun for a specific time and at a certain temperature. Many different conditions have been reported in the literature (see examples in Table 1), and certain groups also used a filter paper or a small platform to prevent the sample from reabsorbing the water during the slowing down of the centrifuge (can take a few min). The latter is very important and will be discussed in the Recommended Method section below. The main advantages of this approach are the speed (a few min to hour), relatively simple to perform provided you have the equipment, and ability to provide a pretty good prediction of the sample behavior during storage/processing. In this case, the researcher also needs to know the history of the sample, pH, and time postmortem. An example of the significant effects of g-force (959, 8,630, and 34,500), centrifugation time (7.5–22.5 min), test temperature (2°C–20°C), and salt concentration (0.0–0.6 M; achieved by adding 8 mL salt solution to 5 g minced meat sample) are shown in Table 2. The information presented demonstrates the importance of selecting the proper conditions to get the best results. Based on the results, the authors recommended using 8,630 g for 7.5 min at 20°C (Zhang et al., 1995). In that experiment, preventing reabsorption of the released liquid was not tested but shown in later studies to be an important factor, as the time required to get the centrifuge to stop can vary depending on the initial speed, type of centrifuge, etc. Therefore, in the Recommended Method (presented below), a screen platform is placed at the bottom of the tube. Another example is using whey protein gels prepared with the same protein concentration but with different ionic strength (Figure 2a). This allowed the researchers to produce gels ranging from clear to opaque gels. The former is clear because they are composed of fine protein strands that allow light to go through, while the latter is composed of protein aggregates that do not allow light to pass through and actually reflect the light, hence appearing as white entities. In terms of WHC, clear gels are known to have a superior WHC compared to opaque gels. The data clearly show how the g force and the presence of a net (added to prevent water reabsorption) are affecting the apparent WHC values and highlight the crucial importance of setting up the method.

Table 2.

Effects of applying different conditions/settings for the centrifugal force test to determine water holding capacity (WHC) of identical meat samples. Data from Zhang et al. (1995).

	Mean value of WHC
		Centrifugal force (g)
Test conditions	Overall	959	8,630	34,500
Test time (min)
7.5	4.0	23.7	4.1	–15.7
15.0	1.1	25.0	–3.7	–17.9
22.5	1.2	27.0	–4.9	–18.5
Test temperature (°C)
2	6.5	28.3	6.0	–14.9
10	1.8	26.6	–3.3	–18.0
20	–1.9	20.7	–7.1	–19.2
Salt concentration (M)
0.0	–2.2	19.3	–4.7	–21.3
0.3	1.9	25.3	–1.9	–17.6
0.6	6.6	31.0	2.1	–13.2
Centrifugal force (g)
959	25.2	–	–	–
8,630	–1.5	–	–	–
34,500	–17.4	–	–	–

Figure 2.

Water holding capacity (WHC) of whey protein isolate gels prepared as clear gels up to aggregated gels (fine strand proteins to aggregated proteins) determined by 3 different methods: (A) centrifugal, (B) filter paper, (C) press method. Note: FP=filter paper. Redrawn from Barbut (1995).

Capillary Force – Filter Paper Method

This method is performed by placing the sample on a filter paper with a specific pore size. The size will affect the absorbing/suction force. Examples of the relationship between pore size and water column height were presented in the Introduction. In this procedure, a specified amount of a meat sample, cut at a certain geometry, is placed on a filter paper, and allowed to stay for a predetermined amount of time. Later, the meat sample is reweighed and the ratio of the meat circle and the formed water ring around it is used to calculate a relative WHC value.

Capillary Suction

A device to measure water suction was also developed by Hofmann (1975) in which a meat sample is placed on a porous gypsum corpus to absorb moisture which displaces air in an overflow pipe connected to the device. The results are expressed as water released per cm² of meat, or per g sample. Figure 2B shows the filter paper results obtained for whey protein isolate (WPI) gels ranging from clear to opaque (very good to very poor WHC, respectively), where time is demonstrated to also be an important factor. Stevenson et al. (2013) used the filter paper method to study WHC of chicken protein and polyacrylamide gels. They compared the results to cooking loss and centrifuge tests as well as pore size measurements obtained by a scanning electron microscope. They indicated that the capillary suction method (placing Watman filter paper # 1 on top of gel samples for 5 d at 20°C) was the most sensitive to detect differences in the gel structure as it is related to WHC. However, this method depends on quite a few variables and is not always suitable for meat samples/ gels with high WHC properties.

Compression Force

Compression force can be applied by simple weights positioned on top of a meat/protein gel sample or using a hydraulic press. This method is quite often cited in the scientific literature. Overall, a specific amount of meat is placed between several layers of filter paper (again to prevent reabsorbing the released liquid when the pressure is removed), and then compressed in between 2 solid flat plates. Samples have been compressed to pressures ranging from a few psi to 2,000 psi for a predetermined time (a few seconds or minutes) as is demonstrated in Table 3. The advantages of this method are its relative simplicity and easiness to perform in a relatively short time. However, it should be recognized that the results are not always well correlated with the drip loss test (gravimetric method). In general, the applied pressure and time should be adjusted to reflect the drip loss test in order to get a relatively good/realistic assessment of WHC. The examples in Table 3 show some of the different test parameters used where settings can vary quite a lot in terms of force, compression time, temperature, etc. Table 4 shows the effect of systematically varying the test parameters when identical meat samples are used. All parameters evaluated (test time, force, salt concentration, and sample weight) significantly affected the results. The authors then calculated the optimal test conditions and recommended sample size of 1 g, compression force of 20 kN, duration of 2 min, and salt (0–2%; note this range also represents industry salt usage).

Table 3.

Examples of different conditions used for the press method reported in the literature.

Investigators	Meat type	Mass (g)	Force reported	Temperature (°C)	Compression time (min)	Salt (%)	Filter paper
Vaisey et al. (1975)	Ground beef	0.5	44.4 kN	*	2	1.3	*
Severini et al. (1984)	Ground pork	0.3	0.39 kN	10	5	0	Used⁎⁎
Suvakov et al. (1984)	Ground pork	0.3	0.39 kN	*	5	4	Used⁎⁎
Min and Ni (1989)	Ground muscle	0.5	0.34 kN	0-4	1	1	Used⁎⁎
Irie et al. (1996)	Beef, pork, chicken, duck	0.5	35 kg/cm2	*	1	0	Toyo #2
Lee et al. (2014)	Minced broiler	0.5	500 psi	*	1	0	Watman #3
Barbera (2019)	Beef and pork	0.25	500 N	2–4	1–10	0	Watman #1
De Sousa Reis et al. (2023)	Minced pork	0.5	10 kg	5	0	0	Watman #1

^⁎Value not reported.

^⁎⁎Type not reported.

Table 4.

Effects of applying different pressures, salt concentration, sample size, and various compression times on the water holding capacity (WHC) results of identical meat samples. Data from Zhang et al. (1993).

	Mean value of WHC (decimal)
		Sample mass (g)
Test conditions	Overall	0.5	1.0	1.5
Test time (min)
1	0.41a	0.32a	0.39a	0.52a
2	0.33b	0.20b	0.33b	0.46b
3	0.28c	0.18c	0.27c	0.40c
Applied force (kN)
10	0.43a	0.38a	0.41a	0.51a
20	0.33b	0.22b	0.32b	0.46b
30	0.26c	0.10c	0.27c	0.42c
Salt concentration (%)
0	0.22c	0.09c	0.21c	0.35c
1	0.37b	0.28b	0.34b	0.49b
2	0.44a	0.32a	0.45a	0.54a
Sample mass (g)
0.5	0.23c	–	–	–
1.0	0.33b	–	–	–
1.5	0.46a	–	–	–

^a-c Averages, within a row and a certain test condition, followed by a diffrent superscript are signigficantly diffrent (p<0.05).

In the example of the whey protein isolate gels, applying relatively low and high pressures resulted in different shapes of the curves (Figure 2C) at the range of 25 to 200 mM sodium chloride (i.e., the clear gel range), but different overall response for the aggregated gels produced with 200 to 500 mM NaCl (the opaque gel range). The again illustrates the importance of employing the right conditions to separate the treatments in a meaningful way.

Application of Heat

Applying heat can be used to evaluate the ability of the meat to maintain its original and/or added water while going through the protein denaturation phase. This is a very important practical measurement for the industry as many meat products are sold as fully cooked items (prepared at the processing plant). Frying, cooking, and smoking lines capable of processing 0.1 to 10 tons/h are common in the industry and being able to measure and predict the WHC is of great importance. The estimates of WHC are often included in the so-called Least-Cost Formulation software programs, and by that allowing the operator to substitute meats and other ingredients (as their cost and availability can change). It should be mentioned that smokehouse yield values can vary depending on the product, casings type and diameter, conditions in the house in terms of relative humidity, air velocity, etc. However, when working with fairly consistent conditions, the experimental WHC values help operate commercial cooking systems. It is also important to collect and report data related to the history of meat (e.g., water loss during storage), otherwise over- or underestimating the WHC can be a problem.

The main advantages of this approach are the direct application to industry conditions and that the test is easy to perform, while the results have a good relationship to the commercial process. One limitation is that just by measuring weight loss in a smokehouse/oven there is no separation of moisture and fat. This can be overcome by using an enclosed test tube system where all the liquids (moisture and fat) are collected and later separated. Therefore, if one runs a parallel cooking loss test in test tubes, it is important to match as many cooking conditions as possible. Overall, this cooking test is reported in many scientific publications and used to compare different meat formulations, and additives (e.g., phosphate, carrageenan, alginate, soy protein isolate) used to enhance yield and texture of meat products.

Comparing Cooking Loss to Other Methods

Some researchers compared methods such as the press method to cooking loss and 3 examples are provided below. Lee et al. (2014) used the filter press method (see conditions in Table 3) and cooking loss of chicken fillets (cooked in covered aluminum pans to 76°C internal temperature) and showed pretty good relationships between the methods.

Zhang et al. (2020) compared broiler meat drip loss results (1 × 1 × 3 cm meat cubes placed in airtight containers and kept at 4°C for 24 hr) to the cooking test results (sealed plastic bags in 80°C water, cooked to 75°C internal temperature). The results showed a similar trend related to the experimental treatments (7.6, 5.1, and 3.6% drip loss vs. 17.4, 15.2, and 12.6% cooking loss for the corresponding treatments).

Cooking loss was also run in parallel to a centrifuge test of fresh bighead carp meat (Liu et al., 2022). For the centrifuge test (see conditions in Table 1), the authors packed the bottom of the centrifuge tube with glass beads to prevent water reabsorption. In the Recommended Method below it is also suggested to use a plastic mesh to achieve the same thing. Overall, the 2 methods showed similar trends (increasing frozen storage time increased moisture loss).

Nondestructive Methods (NMR, Hyperspectral Imaging, Confocal Laser Scanning)

These methods relate WHC to the structure of the muscle/meat proteins and usually employ much more complex scientific equipment. In this article, these methods will only be highlighted briefly with a few examples provided. The reader is encouraged to look for additional information as scientists are currently busy developing inexpensive rapid methods to predict different characteristics of meat (Sánchez et al., 2023). Figure 3 shows, side by side, comparison of the drip loss test, centrifugation test, cooking loss test, low field nuclear magnetic resonance (NMR), and confocal laser scanning microscopy results obtained from the same meat samples. Overall, the authors indicated that when developing nondestructive methods, researchers should compare them to a more traditional/practical method. For example, Yang et al. (2022) used low-field NMR to evaluate sarcoplasmic proteins in pale, soft, exudative (PSE) pig meat and compare to results from a centrifugal test (see conditions in Table 1). They showed similar trends but the NMR analysis helped them more in supporting their scientific hypothesis.

Figure 3.

Effects of aging (1–14 d) on fresh and cooked pork employing (A) the drip loss test according to the Honikel method and a centrifugal test for fresh meat, and the cooking loss test; (B) nuclear magnetic resonance relaxation times; and (C) confocal laser scanning microscopy images. From Straadt et al. (2007) with permission.

Kamruzzaman et al. (2016) compared the results of the traditional drip loss test (2.5 × 2.5 × 2.0 cm red meat samples suspended in plastic jars for 48 h at 4°C) to hyperspectral imaging and reported that selecting 8 wavelengths in the range of 545 to 970 nm can potentially be used to develop an application for estimating WHC of beef, lamb, and pork meat.

Recommended Test Method for WHC

Figure 4 illustrates the recommended test procedure where a centrifuge tube is used to hold the meat while applying 750 g for 10 min at 4°C. The sample (1.5 g) is placed on a screen to prevent reabsorbing the water (during the slowing down of the centrifuge). The test proposed is based on scientific principles (e.g., no permanent sample deformation, no reabsorbing of the water) and is designed to predict WHC of different meat samples/gels. It is important to note that pretty similar test conditions have been already published about 70 years ago (Wierbicki et al., 1957). These conditions have later been used by a number of other research-intensive groups (Hermansson and Lucisano, 1982; Kocher and Foegeding, 1993; see data in Figure 2). The current test parameters have been refined over the years while being applied to different protein systems (e.g., fresh and cooked meat, blood plasma gels, whey protein gels) to assure that, for example, samples are not damaged due to high g force (see above mentioned references for the various conditions tested). However, the test has never caught up as a standard method despite demonstrating its predicting power in different protein gels. It is also understood that some meat samples (e.g., high fat level) might require a different g-force to get the best WHC prediction value. So, it is recommended in such a case to employ both the standard conditions (750 g, 4°C, 10 min) as well as the lab-specific conditions to allow comparing results among different research groups.

Figure 4.

Illustration of the recommended low centrifugal test; parameters provided in the figure. Note: this basic test has also been employed by: Wierbicki et al. (1957) for meat samples; Hermansson and Lucisano (1982) for blood plasma gels; Kocher and Foegeding (1993) for whey protein gels; Barbut (1995) for meat and whey protein gels.

CONCLUSION

Many methods to determine WHC of meat appear in the scientific literature which makes it challenging and sometimes impossible to compare results from different laboratories. This emphasizes the need to select one basic method. This article outlines the main approaches used by meat scientists and concludes by recommending one method and explaining how it was selected.

DISCLOSURES

The author declares no conflicts of interest.