DiscussionTop

Compared with the analysed NDF proportion of substrates (without discounting ashes) that is shown in Table 1, the proportion of cell wall recovery in the extraction process was 0.99, 1.05, 1.09, 0.99, 1.00 and 0.93, respectively. Extraction efficiency was within ± 0.09 units interval, with extreme values for BS and PK. In the former substrate, the lower recovery could be associated to a partial solubilisation of cell wall when processed at a larger scale, as it was also observed by Barrios Urdaneta et al. (2000). In the case of PK, the difference could be due to a noticeable proportion of cracked stones in this by-product that might bias the NDF proportion in the reference analysis from this substrate because of problems in sampling, aspect that should be minimised when a higher initial amount of substrate was used for the large batch extrac-tion respect to chemical determination.

In order to validate the process of cell wall isolation in this experiment, it is assumed that extraction accurate-ly reflects the entire cell wall fraction of all sources, and thus allowed for a homogeneous comparison of their fer-mentation, without interferences from other components of the original feed (Barrios Urdaneta et al., 2000; Zhang et al., 2007). In this regard, it is worth considering that chemical analysis of substrates showed a crude fat con-tent of 88 g/kg DM in PK respect to a range from 3 to 18 g/kg in the other substrates, and a crude protein content of 179, 157 and 133 g/kg DM in SH, PK and DA vs. a range from 48 to 85 g/kg in BP, BS and OH (Table 1).

In other way, some cell wall components such as pectins and to some extent non-lignified xylans are solubilised by neutral detergent (Van Soest, 1994; Jung & Allen, 1995) and thus were not recovered when processed for cell wall extraction. Therefore, fermentation potential of non-pro-cessed BP, which contains from 13 to 20% pectins (Miron et al., 2001; FEDNA, 2019) might be underestimated in substrate comparison. In fact, cell wall fermentation me-asured as gas production after 48 h incubation was 0.86 of that of the whole ingredient of the same batch reported by Ortolani et al. (2020). Other substrates such as SH and DA may also contain noticeable proportions of pectins (around 8 % in both SH and DA; FEDNA, 2012), and thus a reduction in fermentation could be expected when this fraction was removed. However, this might be coun-terbalanced by the protein washout in the cell wall treat-ment, as nitrogen content is negatively correlated with gas production (González-Ronquillo et al., 1998; Cone & Van Gelder, 1999). Thus, despite the 179 and 133 g CP/kg DM of SH and DA (Table 1), fermentation of their extracted cell wall fractions resulted in gas volumes after 48 h close to those reported with the original substrate (Ortolani et al., 2020).

Cell wall polysaccharides from BP and SH were rapid-ly and extensively fermented, at a slower but more cons-tant rate for the latter (Fig. 2), thus making that extent of gas production in the first 24 h interval was 0.74 vs. 0.61 of total gas with BP vs. SH, although both of them reached similar gas volume after 48 h (Fig. 1). Getachew et al. (2004) observed a 0.76 NDF digestibility after 24 h of in vitro incubation. In both cases, such pattern could be related with its high availability of cell wall polysac-charides (Miron et al., 2001; Seo et al., 2009) and low lignin content (0.03 - 0.05 of total NDF). Initially, rate of gas production for PK was relatively constant from 2 to 8 h and then increased up to 12 h, but fermentation from 24 h onwards was notably diminished, and only 0.21 of total gas production was produced thereafter. It seems that availability of fermentable polysaccharides was rapidly reduced, and the masking effect of its high lignin content (0.16 of total NDF) over polysaccharides probably was then more apparent. According to results from Hindle et al. (1995), the proportion of rumen undegradable fraction in palm kernel cake may reach up to 0.37 of total NDF. Non-processed OH had a high cell wall content (Table 1), but in the range previously reported (Garleb et al., 1991; Thompson et al., 2000). The low rate and extent of cell wall fermentation of OH, even lower than that from straw, agrees with previous in situ results (Hsu et al., 1987; Thompson et al., 2000), although it may widely vary de-pending on the oat variety. The low fermentation of OH can be attributed to its high lignin content and the presen-ce of tight lignin/carbohydrate complexes (Garleb et al., 1991). However, for a similar feed such as wheat bran, Miron et al. (2001) suggest that it is cutin (also recovered in the lignin analysis) rather than lignin which restricts microbial fermentation, and Jung & Allen (1995) indicate that determination of lignin as soluble in sulphuric acid tends to underestimate lignin content of grasses.

According to Miron et al. (2001), cell walls from dico-tyledonous substrates (SH, BP, PK and DA) are higher in cellulose and lower in hemicelluloses compared with mo-nocotyledonous cell walls (OH and BS). In any case, pro-portion of cellulose and lignin does not apparently affect the rate or extent of cell wall fraction of these substrates. In fact, correlation of both OMd and 48 h gas production with cellulose, considered as the difference between ADF and lignin, or lignin/cutin contents of incubated cell wall substrates rendered low coefficients (-0.015 and -0.012 for cellulose and -0.325 and -0.433 for Lignin (sa), res-pectively). However, correlations with hemicelluloses, considered as the difference between NDF and ADF, were higher (-0.664 for OMd and -0.685 with 48 h gas produc-tion). Miron et al. (2001) recorded different in vitro micro-bial degradability of cell wall polysaccharides depending on the substrate. Although the potential effect of the lignin proportion is associated with cell wall fermentation pat-tern (Chesson et al., 1983; Thompson et al., 2000), this is not necessarily a direct relationship (Jung & Allen, 1995), as the type of lignin and the extent of linkages between lignin and heteroxylan side-chains, which difficult hemi-celluloses digestion, depend on the nature of the substrate (Jung & Deetz, 1993; Miron et al., 2001).

Methane production can be an index of fermentation efficiency in ruminants, since up to 12% of energy intake is lost as methane (Johnson & Johnson, 1995). Compared to forages, fermentation of some non-forage fibre sources may alter rumen microbiota towards a more amylolytic population, thus leading to a higher propionate produc-tion. Propionate may act as hydrogen sink reducing the utilisation of hydrogen in methanogenesis (Wang et al., 2018). Further, the smaller particle size of these feeds promotes a faster passage rate, which leads to a lower me-thane production (Okine et al., 1989; Beauchemin et al., 2008). This effect is further enhanced as highly digestible fibre sources stay in the rumen for a shorter time, and this may also restrict the time available for fermentation, lea-ding to a lower methane production than less digestible fibre sources. Pardo et al. (2016) estimated a reduction of methane emission from fermentation in dairy goats when increasing proportion of agroindustrial by-products in diet. In general, total methane production (Fig. 3) agreed with gas production pattern (Fig. 1); however, among the highly fermentable sources, from 24 h onwards methane volume from BP was lower than SH, in response to its nu-merically lower methane proportion in total gas (p>0.05). The lack of a specific methane pattern, different to that of total gas production, is expectable considering that no differences were recorded in acetate proportion, and substrates differences in that of butyrate were manifested in lower values for BP and DA, that otherwise rendered a higher VFA production at 12 h. Therefore, qualitatively, fermentation of cell wall fractions from fibrous feeds does not allow for substantial differences in methane production, apart to those expected from the quantitative extent of fermentation, that are manifested in total gas and total VFA production.

However, substantial differences were observed in propionate proportion, which was higher in BP than in PK, OH and DA. It is worth considering that propionate proportion is inversely related with methane (Moss et al., 2000). Having into account that, stoichiometrically, propionate contribution to the volume of gas produced is lower than that from acetate and butyrate (Beuvink & Spoelstra, 1992; Getachew et al., 1997), certain level of underestimation of cell wall fermentation from sugarbeet pulp can be assumed if measured from the total gas production, as it is reflected by its higher total VFA production. However, it was not the case with OMd, which did not differ between BP and SH. Reasons explaining differences in OM that did not match to those in gas production or VFA concentration are not apparent but respond to previously observed non-extracted substrates comparison (Ortolani et al., 2020).

In summary, fermentation of the cell wall fraction of these fibrous feeds is not directly linked to its chemical composition, at least to their cellulose or lignin proportions, although a correlation with their hemicellulose content has been observed. Cell wall of sugarbeet pulp is highly and rapidly fermentable, producing a high proportion of propionate and rendering a low proportion of methane, and that from soybean hulls behaves similarly. The fermentation rate of palm kernel cake was also high during the first 24 h, so a potential contribution can be assumed if included as ingredient in concentrate compound feeds, considering the low rumen retention time of high concentrate diets. Despite structural and chemical differences of cell wall fractions from dehydrated alfalfa and barley straw, their fermentation was very similar in rate and extent.

 

Table 2. MMethane (proportion of total gas) produced for the different substrates (soybean hulls, SH; sugarbeet pulp, BP; palm kernel meal, PK; oat hulls, OH; dehydrated alfalfa hay, DA; barley straw, BS) incubated at successive time intervals, together with organic matter disappearance (OMd) after 48 h.

SEM: standard error of means. Within columns, letters indicate significant differences (p<0.05)

 

 

Figure 3.  Methane production pattern (mmol/g OM) of cell wall fractions extracted from soybean hulls ( ◯ ), sugarbeet pulp (◼ ), palm kernel meal ( ▲ ), oat hulls ( △ ), dehydrated alfalfa hay ( ▢ ) and barley straw ( ● ). Upper bars show standard error of means.

 

Table 3. Total volatile fatty acid (VFA) concentration (mM) and molar proportions (%) of VFA with the different substrates (soybean hulls, SH; sugarbeet pulp, BP; palm kernel meal, PK; oat hulls, OH; dehydrated alfalfa hay, DA; barley straw, BS) at 12 h of incubation.

SEM: standard error of means; BCVFA: branched chain VFA (sum of isobutyrate and isovalerate). Within columns, letters indicate significant differences (p<0.05))