Review: Recent advances in evaluation of bags made from different textiles used in situ ruminal degradation Tiago Neves Pereira Valente1, Edenio Detmann2, and Cla´udia Batista Sampaio2

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1

Instituto Federal Goiano, Posse Campus/GO, Brazil (e-mail: [email protected]); and 2 University Federal of Vic¸osa/MG, Brazil. Received 28 May 2015, accepted 14 July 2015. Published on the web 10 August 2015.

Valente, T. N. P., Detmann, E. and Sampaio, C. B. 2015. Review: Recent advances in evaluation of bags made from different textiles used in situ ruminal degradation. Can. J. Anim. Sci. 95: 493498. Textile bags are used in the laboratory to analyze the indigestible contents (internal markers) of feedstuffs after in situ ruminal incubation. Information is needed on the rate and extent of degradation in the rumen using bags made from different materials. In situ techniques have been used extensively to measure the degradation of feedstuffs in the rumen. However, in situ techniques are prone to variability. This paper reviews the effects of particle size, the material from which bags are made, pore size, tensile strength of the bags, in situ estimation of the levels of indigestible compounds [indigestible dry matter (iDM), indigestible neutral detergent fiber (iNDF), and indigestible acid detergent fiber (iADF)], rumen degradation profiles, and the use of bags made from nylon (50 mm), F57 (Ankom† ), and non-woven textile (100 g m 2). Key words: Internal markers, particle size, ruminal incubations Valente, T. N. P., Detmann, E. et Sampaio, C. B. 2015. Article de synthe`se : Progre`s re´cents dans l’e´valuation des sacs fabrique´s de diffe´rents textiles utilise´s dans la de´gradation ruminale in situ. Can. J. Anim. Sci. 95: 493498. Il y a une excellente occasion d’utiliser les textiles (sacs) dans l’analyse au laboratoire d’aliments, le contenu indigeste (marqueurs internes) du mate´riel apre`s incubation ruminale in situ et la possibilite´ d’utiliser ces diffe´rents sacs pour recueillir des donne´es au sujet des taux et de l’e´tendue de la de´gradation dans le rumen. Les techniques in situ ont e´te´ largement utilise´es pour mesurer la de´gradation d’aliments dans le rumen. Cependant, les techniques in situ sont sujettes a` la variabilite´. Cette revue se penche sur la taille des particules, le mate´riel (tissus) dont est fabrique´ le sac, la taille des pores, la capacite´ de re´sistance a` la traction des sacs et l’estimation in situ des teneurs en compose´s indigestes sous forme de matie`re se`che indigeste (iDM  « indigestible dry matter »), fibre de de´tergent neutre non digestible (iNDF  « indigestible neutral detergent fiber ») et fibre de de´tergent acide non digestible (iADF  « indigestible acid detergent fiber ») ainsi que les profils de de´gradation dans le rumen et l’utilisation des sacs de nylon (50 mm), de F57 (Ankom † ) et de textile non-tisse´ (NWT  « non-woven textile », 100 g m 2). Mots cle´s: Marqueurs internes, taille des particules, incubations ruminales

Knowledge of the levels of neutral detergent fiber (NDF) contents in feedstuff allows the formulation of more adequate diets for farm animals (Knudsen 2001). The method of determining the level of NDF in feed was originally developed by P. J. Van Soest in 1960 and there have been numerous modifications since then. The difficulty in extracting and washing the fibrous residue of some samples complicates the accurate determination of NDF in forage (Mertens 2002). The evaluation of digestion parameters in ruminant animals is assumed to be more accurate when in vivo methods are employed. However, several constraints, such as animal welfare and experimental costs, have been highlighted, and interest in in situ or in vitro evaluations has increased (Broderick and Cochran 2000; Krizsan et al. 2013; Ramin et al. 2013). Among the techniques used, in situ evaluation has been the most recommended (Marinucci et al. 1992; Vanzant et al. 1998) because the measurements are performed in the rumen, where the degradation process is presumed to be more reliable than in vitro (Varel and Kreikemeier 1995; Zhou et al. Can. J. Anim. Sci. (2015) 95: 493498 doi:10.4141/CJAS-2015-100

2012). In addition, in situ evaluation avoids the accumulation of the final products of fermentation, which can affect degradation, as has sometimes been observed under in vitro environments (Cherney et al. 1993). On the other hand, mathematical modeling of in situ degradation profiles allows the estimation of the different parameters of rumen dynamics, including the rate and extent of degradation, the effectively degraded fraction and the rumen fill effect of fiber (Sampaio et al. 2009). The development of the synthetic filter bag system for fiber analyses has created new perspectives for textile utilization in laboratory feed evaluation (Gomes et al. 2011). The filter bag system was found to be more advantageous than the conventional system, which uses crucibles, owing to its lower labor requirement and cost (Cherney 2000). Abbreviations: DM, dry matter; iDM, indigestible dry matter; NDF, neutral detergent fiber; iNDF, indigestible neutral detergent fiber; iADF, indigestible acid detergent fiber; NWT, non-woven textile (100 g m 2) 493

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494 CANADIAN JOURNAL OF ANIMAL SCIENCE

Different textiles, such as nylon (50 mm), F57 (Ankom† ) and non-woven textile (NWT, 100 g m 2), have been suggested to make the bags used in the in situ evaluations, and information regarding the consistency, accuracy, precision of estimates, analytical costs, and operational facilities of these different textiles is needed (Casali et al. 2009). In order to consider a textile useful for ruminal incubation, the bags must have a porosity that prevents the loss of intact or non-degraded particles, while allowing the inflow of rumen fluid and microorganisms and the outflow of degradation products and ensuring that microbial activity inside the bags is similar to that observed in the ruminal environment (Nozie`re and Michalet-Doreau 2000; Ørskov 2000). There is evidence that different textiles can affect the degradation estimates of feedstuffs (Figroid et al. 1972; Van Hellen and Ellis 1977; Weakley et al. 1983; Huhtanen et al. 1994), but information concerning the analytical efficiency of textiles used for in situ evaluations is scarce (Valente et al. 2011a), especially for more recently developed textiles, such as NWT and F57. Photomicrographs of three textiles at 100 magnification are presented in Fig. 1. The superficial structures of F57 and NWT are similar, with a regular geometrical arrangement, and they are less porous than nylon (Fig. 1). In both bags this does not prevent the inflow of microorganisms and the outflow of degraded material, which is necessary for use in rumen studies (Valente et al. 2011a). Particles Sizes The NDF contents of forages and concentrates ground to different particle sizes and using bags made from different textiles are shown in Table 1. Nylon bags produced the smallest NDF concentrations, indicating a loss of particles due to bag porosity. Grinding samples using a 1-mm screen sieve allows efficient extraction of cell contents by the action of neutral detergent, and a greater specific surface area for the action of heat-stable a-amylase. The use of particles ground at 2-mm overestimates NDF contents (Table 1) (Valente et al. 2011b). The lack of standardization constitutes one of the limitations to the correct application of in situ procedures (Vanzant et al. 1998; Broderick and Cochran 2000). Different particle sizes are used in such procedures (Bowman and Firkins 1993; Nocek and Kohn 1988; Kra¨mer et al. 2013). Nocek (1988) suggested using screen sieves of 2 mm and 5 mm for concentrate and forage to compensate the lack of rumination of samples.

Fig. 1. Photomicrographs (100 ) of F57 textile (a), nylon textile (b), and NWT (c) (Valente et al. 2011a).

However, large particles can compromise microbial access to the substrate (Nozie`re and Michaelet-Doreau 2000), and the utilization of 5-mm particles for forage evaluation has not been recommended in more recent approaches [Hvelplund and Weisbjerg 2000; National Research Council (NRC) 2001]. However, the large variability in particle size among different materials seems to be supported by the true variability in the distribution of particle sizes of a milled sample, which can be different for each forage or concentrate (Nozie`re and MichaeletDoreau 2000). When some materials are ground using a 1-mm screen sieve, very thin particles are produced, which can escape through pores in the textile before degradation. This could overestimate the degraded fraction and, consequently, underestimate the non-degraded residue. Despite the heterogeneity among materials, the utilization of particles ground to pass a 2-mm screen sieve seems to favour an equilibrium point between controlling loss of particles and an adequate specific surface for microbial degradation (Krizsan et al. 2015). This approach could be assumed as the current recommendation for in situ procedures of feed evaluation (NRC 2001). Although laboratory extraction with detergents requires 1-mm particles for efficient extraction of soluble compounds, mainly with neutral detergent, this does not compromise the evaluation of fibrous residues obtained with long-term incubations, because significant concentrations of cell contents in the incubation residue are not expected (Valente et al. 2011a). Therefore, extraction with neutral and acid detergents can be done on residues obtained after incubation of 2-mm particles. Tensile Strength The evaluation of tensile strength using a dynamometer showed that the F57 material was not affected by any procedure. The values of tension of rupture were approximately constant after all experimental stages (Table 2). Such patterns can be corroborated by visual examination of the textile. On the other hand, the NWT material showed decreased intension of rupture after ruminal incubation, although the values remained stable after detergent extractions (Valente et al. 2011a). When bags are submitted to long-term ruminal incubation, the textile can lose resistance because the continuous contractions of the rumen submit the bags to constant tension. This decrease in resistance seems to occur despite the absence of alterations in the physical structure of the textile. This shows that although NWT is not altered by chemical extractions with detergents, it can be physically affected by long-term incubation in the rumen. However, this decrease in resistance during ruminal incubation does not seem to cause significant loss of particles because estimates obtained with NWT were similar to those obtained using F57 (Valente et al. 2011b). Nylon presented a lower tension of rupture compared with the other textiles; in addition, it was affected by all experimental procedures, confirming that besides lower resistance, nylon is subjected to damage caused by

VALENTE ET AL. * TEXTILE BAGS FOR RUMINAL DEGRADATION

495

100 Non-degraded fraction of NDF (%)

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90 80

Nylon

70

F57 NWT

60 50 40 30 20 10 0 0

24

48

72

96

120

144 168 192 Incubation time (h)

216

240

264

288

312

Fig. 2. Fitted degradation profiles of neutral detergent fiber (NDF) of coast cross hay using the different textiles (Valente et al. 2011d).

physical, chemical and biological processes, so there is a high probability of rupture during all analytical steps. The gradual decrease in physical resistance culminates in the rupture of fibers and loss of all bags during acid detergent extraction. Similar results were found by Senger et al. (2008), who reported that polyamides are not resistant to hot acid detergent solutions. This can compromise the accuracy of estimates and increases labor because the samples should be taken out of the bags for acid detergent extraction. In situ Estimation of Indigestible Compounds Contents Internal markers are inherent components of feeds that can be used to estimate digesta flow and fecal mass in digestion assays with ruminant animals. Currently, the Table 1. Variation in NDF (percent dry matter) in samples processed at different particle sizes and using bags made of different textiles Textile Item

Corn straw Elephant grass

Corn grain Soybean grain

Particle size (mm)

F57

1 2 Mean 1 2 Mean

90.94 92.14 91.54a 82.63 85.57 84.1

1 2 Mean 1 2 Mean

12.53 23.18 17.86 22.79 36.53 29.66

Nylon NWT

Mean

Forage 87.02 88.9 87.96b 80.59 84.88 82.73

89.91B 91.57A  82.07B 84.73A 

91.77 93.68 92.73a 83 83.77 83.38

Concentrate 12.55 12.25 21.87 22.61 17.21 17.43 22.57 21.79 34.38 33.12 28.48 27.46

12.45B 22.55A  22.39B 34.68A 

CV (%) 1.7 1.7

2.6 7.8

a, b; A, B Means in row followed by different lower case letters within textiles or capital letter for particle sizes are different according to the TukeyKramer test (P B0.01). Adapted from Valente et al. (2011b).

internal markers used most are indigestible dry matter (iDM), indigestible neutral detergent fiber (iNDF), and indigestible acid detergent fiber (iADF) (Detmann et al. 2004). An ideal marker is one that be completely recovered in feces or any segment of the gastrointestinal tract (Owens and Hanson 1992). Without this characteristic, estimates of fecal excretion or digesta flow could be biased. From a theoretical point of view, the capacity to be recovered is inherent to the marker (Detmann et al. 2007); however, indirect influences of the methods applied to estimate its content in samples can apparently bias the estimates of marker recovery. Results for iDM, iNDF and iADF are shown in Table 3. Some authors have suggested using iDM as an internal marker in ruminant nutrition research (Huhtanen et al. 1994), which could be advantageous because of the low analytical cost compared with indigestible fibrous residues. However, the presence of contaminants may compromise the results (Huhtanen et al. 1994; Casali et al. 2009) because detergents are not used to clean the residues after incubation and the withdrawal of microbial debris through anionic action becomes limited (Van Soest 1994). The detergent action could contribute to decrease the contamination by non-microbial residues (feed or rumen content particles) that are normally observed outside bags after removal from the rumen. The contamination by residues in the in situ evaluation of iDM seems not to be constant among different materials (Casali et al. 2009) and possibly is not homogeneous among different samples (Sampaio et al. 2011). Therefore, at least part of the differences among textiles concerning the iDM contents could be confounded with variations in the levels of microbial and non-microbial contaminants. This pattern may cause inconsistencies on inferences and indicates that iDM should not be used as an internal marker. Considering the results of iNDF and iADF, it can be assumed that estimates obtained with both F57 and NWT could be considered equivalent (Table 3). Similar results between

496 CANADIAN JOURNAL OF ANIMAL SCIENCE Table 2. Tension of rupture (kg f) of different textile bags Textile

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Experimental stage Before incubation After ruminal incubation After ruminal incubation and neutral detergent extraction After ruminal incubation and sequential extraction with neutral detergent and acid detergent

F57

NWT

Nylon

30.16 31.47 32.38

35.26 27.38 29.50

24.57 22.36 19.39

31.72

28.02

7.89

Adaptaded from Valente et al. (2011a).

F57 and NWT were reported by Casali et al. (2009) and Valente et al. (2011a), who worked with in situ evaluation of iNDF and laboratory evaluation of NDF, respectively. Krizsan et al. (2015), comparing the concentration of ash-free indigestible neutral detergent fiber, recommend a grind size of 2.0 mm to avoid impaired microbial activity and particle losses. Ruminal Degradation Profiles The non-degraded residues of dry matter (DM) can present variations inherent to the evaluation process. These could be due to a lack of standardization in bag washing procedures after removal from the rumen (Vanzant et al. 1998) or to the presence of microbial contaminants (Huhtanen et al. 1994; Casali et al. 2009), which could be non-homogeneous among replicates (Sampaio et al. 2011). Therefore, the comparison between degradation profiles of DM obtained with different textiles could be imprecise due to the influence of external factors. Such biases are not expected to affect NDF degradation because extraction with anionic detergent withdraws contaminants such as microbial debris (Van Soest 1994). A better understanding of the effects of the

different textiles on DM degradation parameters may be obtained when NDF degradation is simultaneously evaluated. The evaluation of the soluble fraction of NDF presents a different interpretation compared with that done for DM because NDF is insoluble in a neutral medium such as the ruminal environment. Therefore, there is not a true soluble fraction of NDF (Detmann 2010). One of the main constraints to using textiles in the evaluation of insoluble fibrous compounds is the possibility of particle loss during analytical procedures (Hvelpund and Weisbjerg 2000; Casali et al. 2009; Valente et al. 2011c). In this context, when samples are processed to be used in incubation procedures, one must seek a particle size that represents an equilibrium between controlling particles loss and providing an adequate specific surface for microbial degradation (Valente et al. 2011d). The particle size used in this research is that currently recommended for in situ evaluation of feeds (Vanzant et al. 1998; NRC 2001). However, the true particle size after grinding may be different for each material (Nozie`re and Michaelet-Doreau 2000). Therefore, for some materials, grinding to pass through a 2-mm screen sieve can produce particles thin enough to escape through the textile before degradation. This assumption seems to support the variability among feeds concerning loss of fibrous particles. Such a pattern seems to be, at least partially, the cause of the lower estimates of the potentially degradable fraction of NDF when nylon was used, and could directly affect the estimates of DM fractions. The concept of critical time to reach the asymptote was proposed by Casali et al. (2009) as a probabilistic approach to obtain estimates of undegradable fractions by using a single incubation time, noticeably when those fractions are used as internal markers. First, nylon is not recommended for this procedure (Casali et al. 2009; Valente et al. 2011a) because it permits a major loss of fibrous particles. Therefore,

Table 3. Variation in indigestible dry matter, indigestible neutral detergent fiber (iNDF), and indigestible acid detergent fiber (iADF) (percent of dry matter) in samples processed at different particle sizes and using different textile bags Textile Material

F57

Signal grass hay Sugarcane Elephant grass

54.86a 38.08a 37.70a

Signal grass hay Sugarcane Elephant grass

48.65a 33.06a 28.42a

Signal grass hay Sugarcane Elephant grass

31.12 20.21 17.52

Nylon

Particle size (mm) NWT

1

2

Indigestible dry matter (iDM) 35.96b 49.10a 40.82b 45.13b 27.40b 35.59a 32.56 34.63 22.51b 34.63a 27.97b 32.45ab Indigestible neutral detergent fiber (iNDF) 30.91b 44.07a 35.00b 39.88b 24.56b 30.34a 27.27b 30.25ab 17.38b 29.38a 21.06b 25.80ab   

Indigestible acid detergent fibe (iADF) 28.27 24.75b 28.49b 19.54 19.44 19.99 17.71 14.96 18.54

3

CV(%)

53.97a 33.87 34.42a

11.4 9.0 16.5

48.75a 30.44a 28.32a

14.2 7.8 18.7

35.84a 20.19 19.35

14.0 11.9 22.2

a, b Means followed by different letters within textiles or particle sizes are different according to the TukeyKramer test (PB0.05). Adapted from Valente et al. (2011a).

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VALENTE ET AL. * TEXTILE BAGS FOR RUMINAL DEGRADATION

F57 and NWT were compared to evaluate the estimation of undegradable compounds. The average critical times were greater for F57 than for NWT. The maximal estimates of critical times were 249.8 and 219.7 h for DM and 274.5 and 227.7 h for NDF, for F57 and NWT, respectively (Valente et al. 2011d). The maximal estimates of critical times obtained with NWT agree with Casali et al. (2008), who recommended ruminal incubation for 240 h to estimate the undegradable fraction of DM and NDF using NWT bags. However, according to the results of this study, this recommendation seems not to be valid when F57 is used. By considering the maximal estimate of critical time for NDF (274.5 h) and adopting a multiple of 24 h to build an analytical recommendation, at least 288 h (12 d) are necessary to obtain a reliable estimate of the undegradable fraction of NDF, according to the results of Krizsan and Huhtanen (2013) and Kra¨mer et al. (2013). Second, the greater critical times obtained with F57 and NWT compared with nylon were directly caused by lower degradation rates (Fig. 1). Initially, the higher degradation rates obtained with nylon could be attributed to a greater loss of particles during initial incubation times. However, there was little difference between textiles concerning loss of fibrous particles. Therefore, there is not enough evidence to support the loss of particles as the sole or the main cause of differences concerning degradation rates. The structural porosity of NWT and F57 is not regular because the textiles are made by deposition of fibers without weaving, and these textiles show lower porosity than nylon (Casali et al. 2009). Very low porosities can compromise the withdrawal of fermentation gases from inside the bags, which can restrict microbial access to the substrate and, as a consequence, decrease degradation, noticeably at initial incubation times (Ude´n and Van Soest 1984). Such events could lead to lower degradation rates, as shown in Fig. 1 (Valente et al. 2011d). One of the desired characteristics of a textile used to evaluate ruminal degradation is its capacity to allow the inflow of ruminal fluid and microorganisms and at same time allow the outflow of the degraded material (Varvikko and Vanhatalo 1990). Problems concerning this characteristic can affect the slope of degradation curves (Nocek 1988). Decreasing bag porosity can reduce the microorganisms entering the bag (Lindberg et al. 1984) and increase the differences between the microbial populations inside and outside the bags (Nozie`re and Michalet-Doreau 2000). Therefore, the greater critical times obtained with F57 compared with NWT seem to indicate that degradation profiles obtained with F57 could not be completely stabilized. For some forages, the degradation rates were extremely low when F57 and NWT were used. Corrections or adjustments of the degradation profile concerning loss of particles are possible (Nozie`re and MichaletDoreau 2000), but this is not true concerning the underestimation of degradation rate. Nylon is considered to be the standard textile to perform degradation procedures

497

(Hvelplund and Weisbjerg 2000; NRC 2001), and the use of NWT and F57 to estimate parameters of ruminal degradation dynamics is not recommended. Bowman, J. G. P. and Firkins, J. L. 1993. Effects of forage species and particle size on bacterial cellulolytic activity and colonization in situ. J. Anim. Sci. 71: 16231633. Broderick, G. A. and Cochran, R. C. 2000. In vitro and in situ methods for estimating digestibility with reference to protein degradability. Pages 5385 in M. K. Theodorou and J. France, eds. Feeding systems and feed evaluation models. CAB International, Wallingford, UK. Casali, A. O., Detmann, E., Valadares Filho, S. C. et al. 2008. Influence of incubation time and particles size on indigestible compounds contents in cattle feeds and feces obtained by in situ procedures. Braz. J. Anim. Sci. 37: 335342. Casali, A. O., Detmann, E., Valadares Filho, S. C. et al. 2009. Estimation of fibrous compounds contents in ruminant feeds with bags made from different textiles. Braz. J. Anim. Sci. 38: 130138. Cherney, D. J. R. 2000. Characterization of forages by chemical analysis. Pages 281300 in D. I. Givens, E. Owens, and R. F. E. Axford, eds. Forage evaluation in ruminant nutrition. CAB International, Wallingford, UK. Cherney, D. J. R., Siciliano-Jones, J. and Pell, A. N. 1993. Forage in vitro dry matter digestibility as influenced by fiber source in the donor cow diet. J. Anim. Sci. 71: 13351338. Detmann, E. 2010. Fiber in nutrition of dairy heifers. Pages 253302 in E. S. Pereira, P. G. Pimentel, and A. C. Queiroz, eds. Novilhas leiteiras. Graphiti, Fortaleza, Brazil. Detmann, E., Souza, A. L., Garcia, R. et al. 2007. Evaluation of internal markers long term bias in a digestion assay with ruminants. Arquivo Brasileiro de Medicina Veterina´ria e Zootecnia 59: 182188. Detmann, E., Valadares Filho, S. C., Paulino, M. F. et al. 2004. Technical evaluation of the indicators in the estimation of feed intake by ruminants on pasture. Cadernos Te´cnicos de Veterina´ria e Zootecnia 46: 4057. Figroid, W., Hale, W. H. and Theurer, B. 1972. An evaluation of the nylon bag technique for estimating rumen utilization of grains. J. Anim. Sci. 35: 113120. Gomes, D. I., Detmann, E., Valente, T. N. P., Valadares Filho, S. C. and Queiroz, A. C. 2011. Laboratorial evaluation of fibrous compounds in feeds and cattle feces under different physical environments. Arq. Bras. Med. Vet. Zootec. 63: 522525. Huhtanen, P., Kaustell, K. and Jaakkola, S. 1994. The use of internal markers to predict total digestibility and duodenal flow of nutrients in cattle given six different diets. Anim. Feed Sci. Technol. 48: 211227. Hvelplund, T. and Weisbjerg, M. R. 2000. In situ techniques for the estimation of protein degradability and postrumen availability. Pages 233258 in D. I. Givens, E. Owens, and R. F. E. Axford, eds. Forage evaluation in ruminant nutrition. CAB International, Wallingford, UK. Knudsen, K. E. B. 2001. The nutritional significance of ‘‘dietary fibre’’ analyses. Anim. Feed Sci. Technol. 90: 320. Kra¨mer, M., Nørgaard, P., Lund, P. and Weisbjerg, M. R. 2013. Particle size alterations of feedstuffs during in situ neutral detergent fiber incubation. J. Dairy Sci. 96: 46014614. Krizsan, S. J. and Huhtanen, P. 2013. Effect of diet composition and incubation time on feed indigestible neutral detergent fibre concentration in dairy cows. J. Dairy Sci. 96: 17151726.

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