Ruminal and intestinal protein degradability of various seaweed ...

Animal Feed Science and Technology 213 (2016) 44–54

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Ruminal and intestinal protein degradability of various seaweed species measured in situ in dairy cows Usama Tayyab a,∗ , Margarita Novoa-Garrido b , Michael Y. Roleda b , Vibeke Lind b , Martin Riis Weisbjerg a a b

Department of Animal Science, Aarhus University Foulum, Blichers Allé 20, Post Box 50, DK-8830, Tjele, Denmark Norwegian Institute for Bioeconomy Research (NIBIO), Post Box 115, 1431 Ås, Norway

a r t i c l e

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Article history: Received 19 August 2015 Received in revised form 24 November 2015 Accepted 7 January 2016 Keywords: Seaweeds Alternative protein source In situ protein degradability Total tract digestibility Ruminants

a b s t r a c t The use of seaweeds in animal diets is not new. However, little is known about the feed value of seaweed, both in terms of chemical composition and protein digestibility, and regarding variation between species and season. In this study, eight seaweed species of the genus Acrosiphonia, Alaria, Laminaria, Mastocarpus, Palmaria, Pelvetia, Porphyra, and Ulva were sampled in spring (March) and autumn (October and November) 2014 at the coast of Bodø in Northern Norway, and were analysed for chemical composition, in situ rumen degradability and total tract crude protein (CP) digestibility. Ash content in dry matter (DM) was generally high (overall mean 190 g/kg in DM) and varied considerably, between species (P < 0.01) and between seasons (P = 0.02). CP concentration in DM varied both between species (P < 0.0001) and seasons (P < 0.01). Highest CP in DM was found for Porphyra (350 g/kg DM) and lowest for Pelvetia (90 g/kg DM). Spring samples were higher in CP than autumn samples. The effective degradability estimated at 5% rumen passage rate (ED5) of CP varied between species (P < 0.0001) but not between seasons (P = 0.10). The highest ED5 of CP was found for Alaria (550 g/kg CP) and lowest for Ulva (240 g/kg CP). Digestible rumen escape protein (DEP) varied significantly between species (P < 0.0001) but not between seasons (P = 0.06); highest DEP was found for Ulva (530 g/kg CP) and Porphyra (500 g/kg CP). Based on our results, Acrosiphonia, Alaria, Laminaria, Mastocarpus and Palmaria can supply the rumen with high amounts of rumen degradable protein, while Porphyra and Ulva can be used as a source of digestible bypass protein. Pelvetia had a very low degradability and should not be used to feed dairy cows. © 2016 Elsevier B.V. All rights reserved.

1. Introduction The raising demand for both food and feed has increased the search for alternative protein sources. It is no longer environmentally sustainable to increase the land area for cultivation of feedstuffs as this will further contribute to climate

Abbreviations: DM, dry matter; OM, organic matter; N, nitrogen; CP, crude protein; aNDFom, ash free neutral detergent fiber using heat stable amylase; h, hours; min, minutes; a, washout fraction; b, insoluble but potentially degradable fraction; c, rate of degradation of fraction b; ED5, effective degradability calculated at 5% h−1 rumen passage rate; RDP, rumen degradable protein; DEP, digestible escape protein of the total CP intake; SID, digestibility of rumen escape protein in small intestine. ∗ Corresponding author. Present address: Department of Animal Production, Welfare and Veterinary Science, Harper Adams University, TF10 8NB, UK. Fax: +44 1952 814783. E-mail addresses: [email protected] (U. Tayyab), [email protected] (M. Novoa-Garrido), [email protected] (M.Y. Roleda), [email protected] (V. Lind), [email protected] (M.R. Weisbjerg). http://dx.doi.org/10.1016/j.anifeedsci.2016.01.003 0377-8401/© 2016 Elsevier B.V. All rights reserved.

U. Tayyab et al. / Animal Feed Science and Technology 213 (2016) 44–54

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change e.g., slash-and-burn deforestation (Steinfeld et al., 2006). The potential biomass production from the sea has renewed the interest in using seaweeds as animal feedstock. Some seaweeds are protein rich (the CP content can reach up to 470 g/kg DM e.g., Porphyra yezoensis; Arasaki and Arasaki, 1983; Burtin, 2003; Stadtlander et al., 2013) and could potentially be used to decrease the dependency on conventional protein feedstuffs like soybeans. The use of seaweeds in animal feeding is not new; people living on coastal areas have traditionally fed their animals with seaweeds especially during lean feed seasons (Evans and Critchley, 2014). Commercial seaweed production in 2010 reached 19 million tons mainly from seaweed farming in Asia (FAO, 2012); most of which are used for human consumption and industrial purposes, and only a small fraction is used for animal feed (Evans and Critchley, 2014). The Norwegian seaweed industry is reliant on sustainable harvesting of Laminaria hyperborea and Ascophyllum nodosum for alginate production, and for animal food and fertilizer products, respectively (Meland and Rebours, 2012). The increasing demand for seaweed biomass for various applications generated several initiatives to cultivate seaweeds along the cold temperate to the Arctic/Boreal coastal regions of Europe (e.g., Skjermo et al., 2014). In high latitudes, the future ocean warming may be percieved to be favorable as it can increase seaweed growth and production (Krause-Jensen and Duarte, 2014). However, weather anomalies such as increasing storm surges (Roleda and Dethleff, 2011) and extreme high summer temperatures associated with global climate change can contribute to large-scale seaweed dislodgement and disappearance, reduce productivity, and increase blade erosion among different kelps (Roleda and Dethleff, 2011; Roleda, 2015 and references therein). Moreover, seaweeds have complex life cycle that are differentially susceptible to different environmental stressors (Roleda, 2015). For the feed application of seaweed, previous studies focused on the nutritional values of single species or a mixture ˜ of different species fed to small ruminants (Ventura and Castanón, 1998). The nutritive value of seaweeds for ruminants in literature varies widely and it depends on the seaweed and animal species, and the seaweed’s chemical composition. However, in vivo data on protein degradability of seaweeds in dairy cows are scarce (Makkar et al., 2015), to our knowledge only one study examined seaweed protein degradability in the rumen (Gojon-Báez et al., 1998). The aim of this study was to determine the in situ rumen and total tract digestibility of seaweed protein and to evaluate the effect of season and seaweed species on the protein value for dairy cows.

2. Material and methods 2.1. Seaweed collection and sample preparation Eight seaweed species (the brown; Alaria esculenta, Laminaria digitata, Pelvetia canaliculata, the red; Mastocarpus stellatus, Palmaria palmata, Porphyra sp., and the green; Acrosiphonia sp., and Ulva sp.; (hereafter Alaria, Laminaria, Pelvetia, Mastocarpus, Palmaria, Porphyra, Acrosiphonia and Ulva, respectively) were sampled by hand picking in spring (March) and autumn (October and November) 2014 at the coast of Bodø in northern Norway (67◦ 19’00”N, 14◦ 28’60”E at low tide). Ulva was only sampled in autumn. Seaweeds were thoroughly washed and cleaned of sand, epibionts, associated mesograzers and other vertebrates and invertebrates in baths of ambient seawater (salinity 34 mg/L). Clean samples were briefly washed in decreasing salinity of 30% and finally in freshwater; excess water was drained manually and samples were frozen at −20 ◦ C until freeze drying. Dried samples were milled through a 1.5 mm screen with a cutter mill (Fritsch pulverisette 15) for in situ analysis, and a 1 mm screen for chemical analysis.

2.2. Animals The experiment complied with the guidelines set out by the Danish Ministry of Justice with respect to animal experimentation and care of animals used for scientific purposes. Three dry rumen fistulated (#1C, Bar Diamond Inc., Parma, ID, USA) Danish Holstein cows were used for rumen incubations. Cows were fed at maintenance level twice daily with ration consisting of (kg/day as fed) barley straw 2, clover–grass hay 4, concentrate mixture 2.8 and minerals 0.2. Concentrate consisted of (kg/100 kg) Soybean meal 10, barley 40, oat 40, rapeseed meal 3, sugar beet molasses 3 and minerals 4. The ration forage to concentrate ratio was 67:33, and ration concentration of CP and starch was 139 and 137 g/kg DM, respectively. Three duodenally cannulated lactating Danish Holstein dairy cows maintained on a 60:40 forage (grass and maize silage) to concentrate ratio (DM basis) diet were used for intestinal incubations of mobile bags. All cows had free access to fresh drinking water.

2.3. Rumen in situ degradation The seaweed samples (n = 15) were incubated in the rumen in three cows for in situ protein degradation at 8 time intervals (0, 2, 4, 8, 16, 24, 48 and 96 h) using Dacron bags (38 ␮m pore size) according to the standard NorFor procedure (Åkerlind et al., 2011). After rumen incubation, the residues were treated in a Stomacher (Stomacher® 400 Circulator, Seward UK)

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for 5 min with 50–70 mL of distilled water, and thereafter returned to the Dacron bag and washed, before transferred to nitrogen (N) free filter paper (retention value 2, Whatman AGF 607–90 mm) for DM and N analysis. 2.4. Total tract degradation Samples (n = 15) were transferred in mobile bags (12 ␮m pore size), and ruminally pre-incubated in three dry cows for 16 h, each sample replicated twice in each cow. Thereafter the samples were treated with pepsin–HCl and incubated in the small intestine of three lactating cows through the duodenal cannula. The bags were recovered from the feces (Hvelplund and Weisbjerg, 2000). Total tract indigestible CP was estimated as the CP residue in mobile bags after fecal recovery. Rumen degradable, intestinal degradable and indigestible CP was reported as g/kg of original CP seaweed. 2.5. Water soluble fraction The soluble fractions of seaweed samples for both DM and CP were measured by procedure described by Weisbjerg et al. (1990). The milled sample (0.5 g) was weighed out in duplicate in a beaker, and 40 mL distilled water was added. The sample was left for 1 h at normal room temperature. Then the sample was filtrated through N-free filter paper and washed with 4 × 40 mL of demineralised water. 2.6. Chemical analysis Dry matter concentration of seaweed species was estimated as freeze dry matter. Ash was estimated as residue after combustion at 525 ◦ C. The CP values were estimated as N × 6.25 after N analysis by the Kjeldahl method. Neutral detergent fiber (aNDFom) was measured using FibertecTM M6 system (Foss Analytical, Hillerød, Denmark) using heat stable amylase and sodium sulphite according to procedure described by Mertens (2002) and expressed exclusive of residual ash. 2.7. Calculations and statistics 2.7.1. Degradation parameters Degradation profiles of DM and CP were fitted assuming an exponential degradation profile including a lag time using PROC NLIN in SAS (9.4 version, SAS Institute Inc.). Rumen degradable CP was estimated as effective rumen degradability (ED5) at 5% h−1 rumen fractional passage rate (Åkerlind et al., 2011), but also including lag time (Stensig et al., 1994):

ED5 = a + b

c (c + kp)

× (exp (− (c + kp) × lt))

Where a is washout fraction, b is degradable but not soluble fraction, c is fractional rate of degradation, kp is fractional rate of passage (5% h−1 ), and lt is lag time (h). Intestinally degradable CP as proportion of feed CP was estimated as rumen effective escape CP minus indigestible CP. 2.7.2. Statistical analysis All data were analysed using PROC Mixed Model by SAS® 9.4 version (SAS Institute Inc.) with species and season as fixed effects and cow as random effect. Results for species in the text, if not otherwise stated, are presented as average of spring and autumn samples. 3. Results 3.1. Chemical composition The DM concentration of seaweed species ranged between 105 and 283 g/kg among all species sampled across both seasons (Table 1). The highest ash concentration was found for Ulva (483 g/kg DM) and the lowest for Porphyra (128 g/kg DM). Acid insoluble ash was analysed in spring samples, and concentrations were low and below detection level for most samples (results not shown). The CP concentration in DM varied considerable both between species (P < 0.0001) and seasons (P = 0.01) (Table 1). The highest CP in DM was found for Porphyra (347 g/kg DM) and lowest for Pelvetia (90 g/kg DM), but also Acrosiphonia (310 g/kg DM) and Palmaria (223 g/kg DM) were protein rich. The aNDFom concentrations were higher in autumn than spring samples, the highest concentration found for Palmaria autumn sample (501 g/kg DM) and lowest for Alaria (90 g/kg DM) sampled in autumn (Table 1).


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Table 1 Chemical composition of seaweed species. Specie Brown seaweeds Alaria Laminaria Pelvetia Red seaweeds Mastocarpus Palmaria Porphyra Green seaweeds Acrosiphonia Ulva P value

Season

DM

Ash

CP

aNDFom

OMother

Spring Autumn Spring Autumn Spring Autumn

132 237 128 173 229 244

278 139 351 233 219 210

158 127 161 103 105 75

117 90 163 201 293 280

447 644 325 463 383 435

Spring Autumn Spring Autumn Spring Autumn

283 254 160 200 148 105

217 208 165 108 149 107

178 178 257 188 372 321

148 351 421 501 371 408

457 264 157 203 107 164

Spring Autumn Autumn

226 194 143

171 127 483

333 286 162

406 388 286

90 199 69

Species Season

0.09 0.17