Directory UMM :Data Elmu:jurnal:A:Aquaculture:Vol184.Issue1-2.Apr2000:

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Ž .

Aquaculture 184 2000 115–132

www.elsevier.nlrlocateraqua-online

Digestibility of macronutrients, energy and amino

acids, absorption of elements and absence of

intestinal enteritis in Atlantic salmon, Salmo salar,

fed diets with wheat gluten

T. Storebakken

a,)

_{, K.D. Shearer}

_{, G. Baeverfjord}

_,

˚

a b d

B.G. Nielsen , T. Asgard , T. Scott , A. De Laporte

˚

a ₍ ₎

AKVAFORSK Institute of Aquaculture Research A.S. , Sunndalsøra N-6600, Norway

Northwest Fisheries Science Center, NOAArNMFS, 2725 Montlake BouleÕard E., Seattle, WA 98112, USA

BioMar, Myre N-8430, Norway

Amycor, Amylum Group, Burchstraat 10, Aalst B-9300, Belgium Accepted 3 September 1999

Abstract

Ž .

Apparent digestibility coefficients ADCs of macronutrients and energy, and apparent absorp-Ž .

tion coefficients AACs of amino acids and elements were assessed in an experiment with 0.9 kg Atlantic salmon reared in saltwater tanks. Duplicate groups of fish were fed five diets, where 0,

Ž . Ž .

6.25, 12.5, 25 and 50% of crude protein CP from fish meal FM was replaced with CP from Ž .

wheat gluten WG . In Experiment 2, triplicate groups of 0.9 kg salmon were fed a FM diet, a diet Ž .

with 15% of CP from FM replaced with extracted, toasted soybean meal SBM , and a diet with 35% of CP from WG, for 18 weeks. Experiment 2 was designed to determine whether WG caused pathological changes in the intestinal epithelium, and if a diet with 35% of CP from WG could support rapid growth. There was a trend toward increased ADC of fat and energy in the diets with WG, and the diet with 25% WG was ranked significantly higher than the FM control. The ADC of CP and AACs of all amino acids except alanine and lysine increased significantly with increasing proportion of dietary protein from WG, and the results indicate that absorption of individual amino acids from WG was between 94% and 100%. WG is low in lysine, but the results indicate that the requirement for lysine was nearly met and the requirement for other essential amino acids was met even with the highest WG inclusion level, due to high dietary protein concentration and the supplementary amino acid profile of FM. There was no reduced absorption of Ca, P or Mg in the

)_{Corresponding author. Tel.:}_q_{47-71-69-53-14; fax:}_q_{47-71-69-53-01; e-mail:}

[email protected]

Ž .

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salmon fed diets with WG. The absorption of Zn was higher in all the diets with WG than in the FM diet. The fish doubled their weight during Experiment 2, and there were no significant differences in growth among dietary treatments in salmon fed the WG, FM or SBM diets. No indication of intestinal pathology was seen in the salmon fed the FM or WG diets, while 60% of the examined fish fed SBM had SBM-induced changes in the mucosa of the posterior intestine.

Keywords: Wheat gluten–soybean meal–LT fish meal; Digestibility–fat–protein–energy; Absorption–amino acid–elements–minerals–phosphorus; Soybean-induced enteritis; Atlantic salmon — Salmo salar

1. Introduction Ž .

Wheat gluten WG may act both as a source of protein and a pellet binder. Traditionally, starches are used to provide the necessary binding of extruded diets. However, Atlantic salmon has a limited ability to hydrolyse starch in the intestine, and to regulate the blood glucose concentration when the level of digestible carbohydrate is

Ž .

high Hemre et al., 1995a,b . Thus, the amount of starch in diets for salmon is kept low. The use of indigestible hydrocolloid binders used in moist feeds results in reduced

digestibility of both protein and fat Storebakken, 1985; Storebakken and Austreng, .

1987; Fagbenro and Jauncey, 1995; Yamamoto and Akiyama, 1995 . Such problems are Ž

not seen with WG, since it is digestible Fagbenro and Jauncey, 1995; Yamamoto and .

Akiyama, 1995 .

Several experiments have shown that WG can successfully replace a large proportion Ž .

of the fish meal FM in diets for rainbow trout, provided that the diets are supplemented Ž

with lysine, the first limiting amino acid in gluten Pfeffer et al., 1992; Davies et al., .

1997; Schumacher and Wax, 1997 . The literature is contradictory with respect to the

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need for supplementation with arginine Pfeffer et al., 1992; Davies et al., 1997 , which is the second limiting amino acid in gluten. There does not seem to be any need to supplement diets for rainbow trout with threonine, which is the proposed third limiting

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amino acid Pfeffer et al., 1992 . WG is a highly digestible source of protein, as

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demonstrated by Pfeffer et al. 1995 who found an apparent digestibility of 99% for Ž .

crude protein CP when feeding a diet with 92.7% gluten and 1.45% lysine to rainbow trout. The digestibility of protein from gluten was higher than those obtained for various poultry by-products, or hydrothermally treated soy- or field- beans. Similarly, Sugiura et

Ž .

al. 1997 found that WG had the numerically highest value with respect to apparent digestibility of dry matter and protein, when compared with several FMs, poultry by-products, and vegetable protein sources, both in coho salmon and rainbow trout.

Ž .

Skonberg et al. 1998 have shown that incorporation of WG into a diet for rainbow

Ž .

trout did not adversely affect flavour or pigmentation of the fillets. Sugiura et al. 1997 also studied apparent absorption of various elements in WG meal, and found that it was ranked among the best of the ingredients tested both with respect to availability of Ca, Fe, K, Mg, P, Sr and Zn. Other vegetable protein-rich feed ingredients like soybean

Ž .

meal SBM and corn gluten resulted in lower, and partly negative estimates for absorption of several elements in both fish species. The first aim of the present study was to find the digestibility of macronutrients, energy and amino acids, and the

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T. Storebakken et al.rAquaculture 184 2000 115–132 117

absorption of ash and individual elements in diets where up to half of the protein from LT FM was replaced by WG in diets for Atlantic salmon.

Morphological changes are observed in mucosa of the distal intestine of salmonids Ž

when using SBM in the diet van den Ingh et al., 1991; Baeverfjord and Krogdahl, .

1996 . These changes have been suggested to be of a similar nature to gluten intolerance Žceliac disease in man Baeverfjord and Krogdahl, 1996 . The changes can briefly be. Ž .

Ž . Ž .

described as a combination of four features Baeverfjord and Krogdahl, 1996 : 1 a Ž .

shortening of heights of the mucosal foldings; 2 a loss of the normal supranuclear Ž .

vascuolisation of the absorptive cells in the intestinal epithelium; 3 a widening of the central stroma within the mucosal foldings, with increased amounts of connective tissue;

Ž .

and 4 a profound infiltration of inflammatory cells in the lamina propria. Intestinal changes are moderate to small in chinook salmon and rainbow trout when they are fed

Ž .

diets with soy protein concentrate Bureau et al., 1998 . The changes, however, occur when salmonids are fed diets with the alcohol extract obtained by producing the protein

concentrate from soybeans Krogdahl et al., 1995; van den Ingh et al., 1996; Bureau et .

al., 1998 . Soy-protein-concentrate diets supplemented with Quillaja bark sapponines Ž

caused significant intestinal damage in coho salmon and rainbow trout Bureau et al., .

1998 , but purified soy sapponine did not cause soybean-induced enteritis in Atlantic

Ž .

salmon Krogdahl et al., 1995 . Published information concerning the occurrence of such reactions in the intestine of salmonids fed other vegetable protein-rich feed ingredients other than soybean products does not, to our knowledge, exist. The second aim of the study was to find out if similar reactions were seen in the mucosa of the distal intestine of Atlantic salmon fed a diet with WG.

2. Materials and methods 2.1. Experiment 1 — digestibility 2.1.1. Feed ingredients and diets

The macronutrient and amino acid compositions of the feed ingredients containing protein are presented in Table 1, while the element composition of the same ingredients and the calcium phosphate are presented in Table 2. The WG was produced by physical

Ž .

extraction from wheat, dried and finely ground )98% through 160 mm screens . The

Ž .

FM was from herring Clupea harengus , caught and dried less than 2.5 months prior to the experiment. According to the producer, this batch of FM contained 20.5%

water-Ž . y1 y1

soluble CP of total CP ; 0.10 g putrescine kg , 0.45 g cadaverine kg ; -0.1 g histamine kgy1_{, and the protein digestibility assessed with mink was 92.7%. The wheat}

meal was pre-cooked, dried and ground 95% through 160 mm, 99% through 300mm .

screens .

Ž .

Five diets Table 3 were produced with WG replacing 0, 6.25, 12.5, 25 and 50% of the CP from FM. The diets were formulated to contain 440–450 g protein, 320 g fat,

y1 _Ž _.

130 g starch, 77–84 g ash and 15 g phosphorus kg dry matter Table 4 , based on a preliminary analyses of the feed ingredients. The diets were extruded by T. Skretting, Stavanger, Norway, with a diameter of 6 mm.

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Table 1

Macronutrient and amino acid composition of the protein-containing feed ingredients

a b c

WG FM Wheat meal

Ž .

Dry matter DM , g kg 950 928 900

In meal, kg

CP, g 810 748 118

Fat, g 55.6 122.1 11.7

Starch, g 70 0 700

Ash, g 10.1 119 7.1

Gross energy, MJ 22.4 21.0 15.8

Phytic acid, g 2.1 – 0.5

Ž .

Amino acids, g 100 g CP

Asp 3.5 9.1 4.4

Thr 3.7 4.4 2.9

Ser 5.2 4.3 5.1

Glu 38.3 13.0 37.8

Pro 11.4 3.7 12.2

Gly 3.0 5.1 3.4

Ala 2.7 6.2 3.1

Val 3.9 5.0 4.2

Cys 2.2 0.8 2.2

Met 1.6 2.6 1.5

Ile 3.5 4.0 3.6

Leu 7.0 7.3 7.3

Tyr 3.2 2.9 2.7

Phe 5.0 3.6 5.1

Lys 1.5 7.2 2.0

His 1.9 1.9 2.1

Arg 3.6 6.7 3.7

Trp 0.9 1.0 0.9

Amylgluten, Amylum Group, Aalst, Belgium.

Norse LT-94, Norwegian Fish Meal Industries, Bergen, Norway. Used in Experiment 1.

Suprex Wheat, Codrico, Rotterdam, the Netherlands.

The buoyancy of the diets decreased with increasing concentration of WG as the diets with 0–12.5% WG were floating while 25% and 50% WG were sinking pellets. 2.1.2. Fish and facilities

Ž .

Each of the five diets was fed to two groups of Atlantic salmon Salmo salar L.

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with a mean initial weight of 0.94 kg for 2 weeks Experiment 1 . Each group of 20 fish was kept in a 1 m2 _{circular fiberglass tank with a water depth of 45 cm. Each tank was}

Ž y1 _.

supplied with saltwater 32 g l salinity , with a temperature of 7–88C.

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The salmon had been fed a commercial diet T. Skretting, Stavanger, Norway prior to the experiment. During the experiment, the fish were fed in excess, with electrically

Ž . y1

driven disc feeders Akvaprodukter, Sunndalsøra, Norway , every 10 min, 7 h day . Excess feeding was verified by uneaten feed trapped by strainers in water outlet from the tanks, but feed intake was not quantified. At the termination of working hours, feed floating in the tanks was removed and the fish were left without feed until next morning

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T. Storebakken et al.rAquaculture 184 2000 115–132 119

Table 2

Mineral composition of the protein-containing feed ingredients and the calcium phosphate

a b c d

WG FM Wheat meal Dicalcium phosphate

Ž .

Elements, kg DM

Ca, g 1.0 23.9 0.1 305

Cu, mg 6.26 3.26 0.82 12.0

Fe, mg 77.7 155 9.6 3347

K, g 1.3 12.2 1.0 2.3

Mg, g 0.70 2.64 0.15 6.13

Mn, mg 35.4 5.0 3.6 457

Na, g 0.26 14.5 0.10 2.6

P, g 2.5 22.1 1.2 205

Sr, mg 2.3 43.3 0.9 198

Zn, mg 59.6 121.4 10.0 242

a – c

See footnotes to Table 1.

Feed grade, Windmill Dicalpos, The Windmill Feed Phosphates, Tessenderjo Chemie, Rotterdam, the Netherlands.

in order to avoid ingestion of feed that may have leached nutrients. When floating feed was present in one or more tanks, the removal procedure was done in all tanks, in order to standardise the stress on the fish.

2.1.3. Sampling and analyses

Faeces were collected by careful stripping at 10 and 14 days of feeding. Stripping of

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faeces was carried out as described by Austreng 1978 . Prior to stripping, the fish were anesthetised with metocain, 60 mg ly1_{, dissolved in saltwater.}

Table 3

Formulation of the diets for Experiment 1

Protein from gluten, percentage of total protein 0 6.25 12.5 25 50

Formulation, g kg diet

WG 0 36.8 73.4 146.3 291.3

FM 567.9 530.1 492.6 418.3 269.9

Wheat meal 174.0 169.9 165.7 157.4 141.0

Fish oil 239.5 241.1 242.9 246.3 253.2

Inert marker, Y O2 3 0.1 0.1 0.1 0.1 0.1

Dicalcium phosphate 13.4 16.9 20.2 26.5 39.4

Vitamin and mineral premix 5.2 5.2 5.2 5.2 5.2

See footnotes to Tables 1 and 2.

Denofa, Fredrikstad, Norway.

Sigma, St. Louis, MO, USA.

Active ingredient supplied per kilogram feed. Vitamins: retinol acetate, 5 mg; vitamin D , 4.8 mg;3

a-tocopheryl acetate, 400 mg; thiamin–Cl, 15.3 mg; riboflavin, 26 mg; pyridoxine–Cl, 15.3 mg; Ca-pan-thotenate, 88.9 mg; niacin, 153.1 mg; folic acid, 5.2 mg; vitamin B , 20.0 mg; biotin, 150.0 mg; myo-inositol,12

408.2 mg; vitamin K , 40.0 mg; ascorbate polyphosphate, 1 g all vitamins were from F. Hoffmann La Roche,3

Basel, Switzerland ; choline chloride, 2.4 g. Minerals: 48.0 mg CuSO ; 2.0 mg KI; 109.4 mg MnSO ; 0.4 mg4 4

Ž .

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Table 4

Chemical composition of the diets in Experiment 1

Protein from gluten, % 0 6.25 12.5 25 50

Ž .

Dry matter DM , g kg 942 956 947 952 951

In diet, kg

CP, g 431 438 437 450 458

Fat, g 319 323 322 322 315

Starch, g 116 106 114 114 114

Ash, g 80.5 78.5 76.1 75.6 71.5

Gross energy, MJ 24.1 24.4 24.3 24.4 24.6

a a

Phytic acid, g ND ND 0.1 0.3 0.5

Ž .

Amino acids, g 100 g CP

Asp 7.8 6.8 7.2 6.5 5.4

Thr 4.0 3.5 3.8 3.5 3.2

Ser 3.8 3.6 4.0 4.1 4.1

Glu 13.3 13.7 16.4 19.4 23.7

Pro 4.1 4.3 5.2 6.2 7.8

Gly 4.7 4.1 4.3 4.1 3.7

Ala 5.5 4.7 5.0 4.5 3.8

Val 4.7 4.3 4.6 4.4 4.1

Cys 0.8 0.9 1.0 1.2 1.4

Met 2.3 2.1 2.2 2.0 1.8

Ile 3.9 3.5 3.8 3.7 3.6

Leu 6.8 6.2 6.8 6.7 6.5

Tyr 2.7 2.4 2.7 2.8 2.8

Phe 3.5 3.2 3.7 3.9 4.1

Lys 6.4 5.4 5.6 4.7 3.6

His 1.8 1.7 1.8 1.7 1.8

Arg 6.2 5.6 5.8 5.3 4.6

Trp 0.9 0.9 0.9 0.9 0.8

Ž .

Elements, kg DM

Ca, g 17.7 17.5 17.6 18.2 19.0

Cu, mg 16.9 17.1 17.4 17.7 16.6

Fe, mg 143 148 159 176 206

K, g 7.81 7.37 6.63 5.66 4.41

Mg, g 1.66 1.58 1.51 1.41 1.23

Mn, mg 52.2 59.1 58.7 63.2 67.7

Na, g 7.89 7.32 6.95 6.09 4.21

P, g 15.5 15.3 15.1 15.2 15.1

Sr, mg 27.2 25.8 24.7 23.4 21.1

Zn, mg 169 160 157 159 153

Below detection limit.

The faeces were pooled from each tank, freeze-dried, and ground with a pestle and Ž mortar. Feed ingredients, diets and faeces were analysed for nutrient composition dry

matter, CP, crude fat, starch, energy, ash and concentration of yttrium oxide as

Ž .

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T. Storebakken et al.rAquaculture 184 2000 115–132 121

Ž . Ž .

to Shearer 1984 . Phytic acid was analysed according to Thomson and Erdman 1982 ,

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following the experimental protocol described by Stone et al. 1984 . Amino acids were

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analysed according to Anonymous 1998 . 2.1.4. Calculations and statistical analyses

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Apparent digestibility coefficients ADC of macronutrients and energy, and apparent

Ž .

absorption coefficient AAC of amino acids, ash and elements were calculated as

Ž .

described by Austreng 1978 , except that Y O was used as inert marker. The apparent2 3

AACs for several elements, such as Ca, Mg, Na, and Sr, are underestimated due to the Ž

uptake of elements from the water and excretion by the intestine Storebakken et al., .

1998 .

The results were analysed statistically by one-way analysis of variance with tank

Ž .

mean as the unit of observation. Significant P-0.05 differences were ranked with Duncan’s multiple range test. Results concerning the ADC of protein and AACs of amino acids were analysed by regression according to the following model:

ADC or AACsaqbG, G is protein % of total CP from WG.

Ž

.

2.2. Experiment 2 — histology in the distal intestine and growth 2.2.1. Feed ingredients and diets

Three diets FM control; SBM control diet with 15% CP from SBM; WG diet with

Ž . Ž .

35% of CP from WG the same type as used in Experiment 1 Table 5 were

Table 5

Formulation and chemical composition of the diets for Experiment 2

Diet name FM SBM WG

y1 Formulation, g kg diet

WG 0 0 167

FM 507 434 321

SBM 0 127 0

Wheat meal 193 134 197

Fish oil 295 300 310

Constant ingredients 5 5 5

y1 Chemical composition, kg diet

Ž .

Dry matter DM , g 932 932 942

CP, g 413 396 395

Fat, g 341 330 347

Ash, g 60 61 41

Gross energy, MJ 25.3 24.9 25.8

See footnote to Table 1.

Norse LT-94 from Herring, Norsildmel.

DenoSoy, extracted, toasted, with hulls, Denofa, Fredrikstad, Norway.

NorSalmOil, Nordsildmel.

e_{Vitamin premix, 2 g kg}y1_{; mineral premix, 2 g kg}y1 _Ž_{formulations proprietary to BioMar ; Carophyll}_.

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formulated and extruded to 9 mm pellets by BioMar, Myre, Norway. The level of SBM inclusion was selected to facilitate rapid growth, but still enabling the detection of histological changes in the intestine. The FM, SBM and WG were from the same producers as in Experiment 1. The FM was from herring, from a different batch than in

Ž y1

Experiment 1 74.5% CP; 21.9% water-soluble CP; 0.66 g cadaverine kg , 0.16 g

y1 y1 _.

NH -N kg₃ ; 0.29 g histamine kg ; 92.1% mink protein digestibility . 2.2.2. Fish, facilities and statistical analyses

The experiment was carried out at AKVAFORSK, Averøy, Norway, with Atlantic salmon with an initial weight of 0.95 kg, for 18 weeks. Each diet was fed to apparent satiation to salmon in three floating net-pens with a volume of 125 m3_{, initially with 350}

salmon in each pen. During the first month, the fish were fed by hand, one meal a day. During the rest of the experiment, the fish were fed one meal by hand and one by electric feeders. The salinity of the water was 33 g ly1_{. The temperature at 3 m was 5}₈_C

at the start, rapidly declined to 3.0–3.58C, and then increased to 12.58C at the end of the experiment, with an average temperature of 7.48C. The oxygen concentration in the pens, measured daily, was never below 8 mg ly1.

The growth results were statistically analysed by one-way ANOVA as described for Experiment 1.

2.2.3. Histology

Five fish from each of the two control diets and 15 salmon fed the diet with WG were obtained for examination of intestinal morphology. The lower number of samples from the control diets was chosen because the lack of reaction to FM and the development of

intestinal pathologies in fish fed SBM have been described previously Baeverfjord and .

Krogdahl, 1996 . The fish were killed individually by an overdose of metacain. The fish were not fasted before sampling. The abdominal cavity was opened, the distal intestine was identified and dissected, and a 1-cm long piece was cut from the middle of the distal intestine. The ring of intestinal tissue was cut open, and the sample was rinsed in saline Ž_{9 g l}y1_. _{to remove the gut contents. The sample was fixed by immersion in}

Ž .

phosphate-buffered formalin 4%, pH 7 and stored in a refrigerator. After fixation, the samples were dehydrated in ethanol, equilibrated in xylene, and embedded in paraffin according to routine techniques. Sections of approximately 5mm were cut and stained

Ž .

with haematoxylin and eosin H & E .

Intestinal mucosa were classified into one of three categories based on a combination

Ž . Ž .

of four features Baeverfjord and Krogdahl, 1996 : 1 a shortening of heights of the

Table 6

Ž .

ADCs for fat, starch and energy mean"S.E.M. , ns2 tanks per feed

Protein from wheat gluten, % 0 6.25 12.5 25 50

ADC, %

a,b a,b,c a c b,c

Fat 91.6"0.6 92.1"0.3 90.9"0.2 93.1"0.4 92.6"0.2 Starch 72.2"3.9 63.5"1.1 64.0"0.5 66.0"1.4 61.2"1.4

a,b b a c c

Energy 87.0"0.4 87.3"0.3 86.1"0.4 88.5"0.2 88.5"0.2

a,b,c

Ž .

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Storebakken

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Aquaculture

184

2000

115

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Table 7

Ž .1

ADCs for CP and AAC of individual amino acids mean"S.E.M. , ns2 tanks per feed

2 _Ž _. 3

Dietary protein 0 6.25 12.5 25 50 Regression r P slope)0 Estimated ADC or

from WG, % AAC in gluten, %

ADC, %

a a a b b

Protein 88.6"0.4 89.7"0.1 89.7"0.4 92.2"0.6 93.6"0.1 88.8q0.118G 0.94 0.0002 100.6 AAC, %

Asp 87.9"1.4 88.7"1.4 88.3"0.2 89.2"0.8 90.0"0.4 87.9q0.058G 0.72 0.018 93.7

a a,b a a b

Thr 92.9"0.5 93.8"0.5 93.3"0.1 93.3"0.3 95.0"0.2 93.0q0.035G 0.76 0.010 96.5

a a a a b

Ser 92.1"0.7 93.3"0.2 93.4"0.1 93.1"1.0 95.9"0.2 92.3q0.065G 0.84 0.0024 98.8

b c c a d

Glu 95.5"0.3 96.6"0.1 96.3"0.0 94.2"0.3 98.3"0.1 95.8q0.053G 0.94 0.0001 99.4

a b c d e

Pro 92.9"0.1 94.5"0.3 95.2"0.2 96.6"0.0 97.8"0.2 93.7q0.090G 0.95 0.0001 102.7

a a a,b b c

Gly 88.3"0.4 88.7"0.5 89.6"0.2 90.8"0.6 92.8"0.4 88.3q0.092G 0.97 0.0001 97.5 Ala 92.6"1.5 94.1"0.1 93.8"0.3 93.9"0.6 94.7"0.2 93.2q0.030G 0.53 0.11 96.2 Val 94.9"0.5 95.6"0.3 95.1"0.1 95.5"0.3 96.6"0.1 95.0q0.029G 0.79 0.0060 97.9

a b b c d

Cys 80.4"0.6 83.4"0.7 84.6"0.4 89.2"0.1 92.3"0.2 81.6q0.233G 0.96 0.0001 104.9 Met 92.8"0.4 93.7"0.9 93.3"0.2 93.9"0.4 95.6"0.3 92.9q0.052G 0.86 0.0016 98.1

a a a a,b b

Ile 95.6"0.5 96.1"0.1 95.6"0.2 96.1"0.1 97.0"0.2 95.6q0.026G 0.83 0.0031 97.8

a a a a b

Leu 96.1"0.4 96.6"0.1 96.3"0.2 96.7"0.1 97.5"0.1 96.1q0.027G 0.89 0.0005 98.8 Tyr 95.3"0.7 95.7"0.1 95.8"0.2 95.9"0.2 97.2"0.2 95.3q0.035G 0.86 0.0013 98.8

a a,b a,b b c

Phe 95.2"0.6 95.7"0.0 95.8"0.2 96.3"0.2 97.4"0.1 95.3q0.041G 0.93 0.0001 99.4 Lys 96.1"0.3 96.3"0.1 96.0"0.1 95.8"0.1 95.8"0.3 96.1–0.008G 0.69 0.069 95.3

a a a a b

His 93.8"0.8 94.7"0.1 94.7"0.1 94.9"0.1 96.3"0.3 94.0q0.043G 0.86 0.0088 98.3

a a,b b b c

Arg 96.3"0.1 96.6"0.2 96.8"0.1 96.9"0.1 97.4"0.1 96.4q0.021G 0.93 0.0001 98.5

a a a b b

Trp 87.8"0.7 87.7"0.5 88.4"0.1 91.2"1.1 92.1"0.2 87.6q0.099G 0.89 0.0005 97.5

1_{Different superscripts}a,b,c,d,e_{indicate significant P}_Ž _-_{0.05 differences among diet means.}_.

2 _Ž _.

ADC of protein or AAC of amino acidsaqbG, G is protein % of total crude protein from WG.

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Ž .

mucosal foldings; 2 a loss of the normal supranuclear vascuolisation of the absorptive Ž .

cells in the intestinal epithelium; 3 a widening of the central stroma within the mucosal Ž .

foldings, with increased amounts of connective tissue; and 4 a profound infiltration of inflammatory cells in the lamina propria. The results were categorised as follows:

0: No deviations from normal morphology;

A: Intestinal tissue shows changes which are typical for diets containing solvent-ex-tracted SBM, as described above; and

B: Morphological changes observed, but to a lesser extent than in A andror not all criteria present.

3. Results

y1 _Ž _.

The WG contained 2.1 g phytic acid kg Table 1 . Thus, phytate-P only accounted

Ž .

for 26% of total P in the gluten Table 2 . Relative to the herring meal, gluten contained more glutamic acid, phenylalanine, proline, tyrosine and serine, while the content of

Ž .

aspartate, glycine, alanine, arginine, valine, methionine and lysine was lower Table 1 . The amino acid composition of the wheat matched that of the WG. Compared with the FM, WG contained considerably less Ca, K, Na, P and Sr, moderately less Fe, Mg and

Ž .

Zn, and more Cu and Mn Table 2 .

Ž .

The chemical composition of the diets Table 4 was characterised by a slight increase of protein, from 431 to 458 g kgy1, with increasing proportion of protein from WG. The amino acid composition of the diets varied according to the composition in the feed ingredients. Dietary fat, starch and energy were similar in all diets. Phytic acid was not detectable in the FM control diet or in the diet with 6.25% gluten, and it increased with increasing gluten inclusion up to 50%. The levels of Ca, Fe and Mn increased in the diets with increasing gluten content, Mg, Sr and Zn decreased slightly, while the decrease in K and Na was of a higher magnitude. The concentrations of Cu and P were similar in all five diets.

Table 8

Ž .

Apparent absorption of elements mean"S.E.M.

Protein from wheat gluten, % 0 6.25 12.5 25 50

Apparent absorption, %

Ca y35"5 y39"1 y31"8 y33"2 y29"4 Cu 37"2 41"2 47"4 47"2 47"5 Fe 11.6"1.0 10.6"0.8 11.0"0.2 13.6"6.2 14.2"0.6 K 96.0"0.1 93.3"0.6 93.3"1.2 93.8"1.3 94.6"1.2 Mg y500"76 y546"15 y537"44 y517"31 y451"6 Mn y1"8 y1"2 5"3 7"1 10"1 Na 41.0"1 29.6"7 5.7"25 20.3"16 26.6"20 P 35.7"0.7 32.6"0.0 34.4"0.8 34.1"4.9 39.3"1.3 Sr y382"88 y434"1 y444"9 y442"5 y397"8

a b b b b

Zn 33.5"3.3 45.4"1.6 46.9"4.7 48.2"0.2 45.3"1.6

a,b _Ž _.

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( )

T. Storebakken et al.rAquaculture 184 2000 115–132 125

Table 9

Ž .a

Weight and growth of the salmon in Experiment 2 mean"S.E.M. , ns3 pens per feed

Diet name FM SBM WG

Fish weight, g

0 weeks 952"14 953"10 956"11

18 weeks 2186"28 2073"27 2176"49

SGR 0.66"0.02 0.62"0.02 0.65"0.01

Ž .

No significant P)0.05 differences were present.

b_{Specific growth rate, 100 d}y1 _w_{ln W}_Ž _._y_{ln W}_Ž _._x_{; d, feeding days; W, mean fish weight in a pen;} F I

subscripts I and F, initial and final values.

Ž .

The ADC for fat Table 6 was significantly higher in the diet with 25% gluten than in the FM control diet and the diet with 12.5% gluten, while it did not differ from the

Ž .

other two diets. There also was a tendency Ps0.077 that the ADC of starch was higher in the FM control diet than in the diets with gluten. The ADC of energy was highest for the two highest inclusion levels of gluten, and it was higher in the diet with 6.25% gluten than in the diet with 12.5% gluten.

Ž .

The ADC of CP and AACs of all amino acids Table 7 , except alanine and lysine, showed a significant increase with increasing proportion of protein from WG. The AAC

Ž .

of lysine was negatively correlated rs0.72 to dietary concentration of WG. The ADCs of protein for the diets with 25% and 50% WG were significantly higher than the values obtained for the diets with less WG.

Ž .

The only significant difference in apparent absorption of elements Table 8 was that Zn was higher in all the diets with WG than in the FM control diet. The absorption estimates for Ca, Mg and Sr were negative. The estimate for absorption of Na was biased by a large non-systematic variation. The absorption of P was not significantly different for the various diets, and the mean absorption of P was 35%.

Ž .

The fish doubled their weight during Experiment 2 Table 9 , and there were no significant differences among the various dietary treatments in fish weights or growth. Results from the histological evaluation of the distal intestine of the salmon are presented in Table 10. Fish fed the diet with FM as the only source of protein displayed no changes in intestinal tissue. In the fish fed the diet with 15% of protein from SBM, three of the five salmon displayed changes which were typical for diets containing

Table 10

Pathological changes of the distal intestine of salmon fed a FM control diet, a diet with 15% of the protein from extracted, toasted SBM, and a diet with 35% protein from WG

a a

Diet Number of samples Observations A B

FM 5 No changes from normal morphology 0r5 0r5

SBM 5 Variable changes 1r5 2r5

WG 15 No changes in 14 of 15 evaluated fish. 0r15 1r15

Minor changes in 1 of 15 fish

A: Typical changes as described for solvent-extracted SBM. All criteria fulfilled. B: Pathological changes indicated, but less grave or not all criteria present.

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solvent extracted SBM. For this group as a whole, the changes were somewhat less

Ž .

severe than observed previously Baeverfjord and Krogdahl, 1996 , but the effect was nevertheless clearly identified. In salmon fed the diet with 35% protein from WG, one fish of the 15 which were examined showed very moderate reactions. The changes consisted of inflammatory cell infiltration of submucosal connective tissue structures, and were of unspecific nature.

4. Discussion

The low amount of phytic acid and the low proportion of phytic P in the gluten diets indicate that endogenous phytases were active during processing of the feed ingredient,

since about two thirds of the P in cereal grains is in the form of phytate Pointillard, .

1993 . Phytase from wheat bran has previously been used to hydrolyse phytic acid from

Ž .

cannola meal in blends of fish silage, cannola meal and wheat bran Stone et al., 1984 and phytase from wheat middlings has been shown to increase P bioavailability in diets

Ž .

for pigs Lei et al., 1997 .

The slight increase in dietary CP content, from 430 to 458 g kgy1 diet, with increasing content of WG is a direct result of the diet formulation, aimed at keeping the diets iso-energetic, and with a constant concentration of starch. Mundheim and Opstvedt Ž1990 did not find significant effects on ADC of protein by replacing protein with fat,.

while dietary starch affects ADC of other nutrients in Atlantic salmon Aksnes, 1995; .

Hemre et al., 1995a; Grisdale-Helland and Helland, 1997 . The increased Ca concentra-tion in the diets with increasing gluten inclusion is a result of keeping the concentraconcentra-tion of P constant, by supplementation with calcium phosphate, while the variation in the concentration of the other elements is a result mainly of differences between the FM and gluten. The differences in elemental concentration, however, were small to moderate, and should not affect uptake of the actual elements or interact with the uptake of other elements.

The growth rate obtained in Experiment 2 by feeding the diet with 35% protein from Ž

WG and the FM diet was similar, on a thermal growth coefficient Iwamata and Tautz, .

1981 basis, to that observed when feeding a diet similar to the FM control to 1 kg

Ž .

salmon at higher water temperatures Einen and Roem, 1997 . This high growth rate illustrates that WG may successfully replace one third of the FM protein in high-energy diets for salmon.

The digestibility of fat and energy was comparable to, or higher than, that previously Ž

reported for Atlantic salmon fed extruded high energy diets in saltwater Johnsen et al., .

1993; Aksnes, 1995; Thodesen and Storebakken, 1998 . In contrast to what has been

Ž .

reported when replacing FM with SBM Refstie et al., 1998 , replacement of LT FM with WG did not result in reduced ADC of fat or energy. On the contrary, ADC of fat was higher in the diet with 25% of the protein from gluten than that of the FM diet. The higher ADC of energy in the diets with 25% and 50% gluten than the FM control diet is the sum of effects of ADC of fat and protein, modified by the tendency toward lower ADC of starch in the diets with WG. This tendency toward reduced ADC of starch may

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( )

T. Storebakken et al.rAquaculture 184 2000 115–132 127

Ž .

be ascribed to the a-amylase inhibitor found in wheat Hofer and Sturmbauer, 1985 . This inhibitor is a protein, which may be concentrated in the protein fraction when producing the gluten.

The ADC of protein in the FM control diet agrees with previous findings with LT-94 Ž

FM in extruded high-energy diets fed to Atlantic salmon in saltwater Johnsen et al., .

1993; Aksnes, 1995 . The AAC of amino acids was generally 2–3% units higher than the results reported for a cold-pelleted LT-70 FM-based diet with approximately 180 g

y1 _Ž _.

fat kg Skrede et al., 1998 . This illustrates that the quality of the herring meal used in the diets was high. In spite of this, the results showed that the majority of amino acids were better digested in gluten than in LT herring meal. Consequently, the ADC of CP increased with increasing inclusion of WG in the feed. The regression equation in Table 7 indicates that when WG is 100% of dietary CP, it is predicted that several of the amino acids will be completely absorbed from the diet. However, such extrapolations should be made with care, since the highest level of inclusion used in the experiment

Ž Ž 2_..

was 50% of the protein from WG, and that 12% 100= 1yr of the variation in ADC of protein was due to other factors than the diet, or interactions among dietary components. The estimated ADC of CP, using the regression in Table 7, for a diet with WG as the only source of protein was 100%, in keeping with the value of 99% obtained

Ž .

by Pfeffer et al. 1995 in rainbow trout. The ADC of protein for the diet without gluten was 88.8%.

The digestabilities presented in this paper were apparent values, not corrected for endogenous losses. True digestabilities of protein and amino acids are higher than the corresponding apparent values, because of the endogenous cost of protein digestion ŽBatterham, 1994 . It is probable that the tendency of lowered AAC of lysine with. increasing dietary gluten level is explained by the endogenous lysine excretion repre-senting a relatively higher proportion of faecal amino acids with increasing dietary gluten levels.

Availability of cysteine in feed ingredients for salmonids may be low Skrede et al., .

1981, 1998 due to the negative effects of heat when the feed ingredient is dried, such as

Ž .

denaturation, oxidation and cross-linkage Opstvedt et al., 1984 . Furthermore,

sulphur-Ž .

containing amino acids are limiting in SBM Lovell, 1989 . Thus, the combination of high concentration and availability of cysteine in WG may contribute to an improved balance of absorbed amino acids in salmonids fed diets with limited availability of cysteine or ingredients which are low in sulphur-containing amino acids.

There is limited information available on the amino acid requirement of Atlantic salmon reared in saltwater, but the requirement of lysine has recently been established to 16–18 g kgy1 dietary dry matter or 0.79–0.89 g MJy1 digestible energy in slowly

Ž .

growing salmon with an initial weight of 0.4 kg Berge et al., 1998 . These workers observed mortality and poor growth in the beginning of the study, and SGR was only 0.69 at 88C in the best group at the end of the experiment. When compared to

Ž .

whole-body amino acid profiles in Atlantic salmon Wilson and Cowey, 1985 , lysine is

Ž .

the first limiting amino acid in WG. Wilson and Cowey 1985 reported that whole salmon contained as much as 9.3 g lysiner100 g amino acids, while the lysine concentration in WG is only 1.5 gr100 g CP. This illustrates the need for lysine supplementation to the diets when a major amount of the protein originates from WG.

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The optimal supplementation of lysine and eventually other essential amino acids must be resolved in future experiments with Atlantic salmon.

In spite of the imbalance in amino acid composition of the gluten, the good growth obtained by replacing 35% of FM protein by WG in Experiment 2 indicates that the essential amino acid supply for protein synthesis may have been adequate. This seems to also be the case for the diet with 50% of CP from gluten. The lysine supply from this diet may be estimated based on the following assumptions: 1 kg of gain in a 1-kg

Ž .

Atlantic salmon constitutes of 180 g CP Shearer et al., 1994 . Whole-body protein of

Ž .

Atlantic salmon contains 9.28 g lysiner100 g amino acids Wilson and Cowey, 1985 . If

Ž .

the amino acid-N accounts for 92% of CP Storebakken, unpublished , the net lysine requirement for growth, thus, should be approximately 15.4 g kgy1 _{gain. The lysine}

content of the diet with 50% protein from WG was 3.6 gr100 g CP, the dietary CP concentration was 458 g kgy1_{, and AAC of lysine was 95.8. If assuming a feed}

conversion ratio of 1 kg dry feed intake per kilogram gain, the digestible lysine intake would be 15.8 g kgy1

gain. The gross lysine content in the diet with 50% of CP from WG, not corrected for digestibility, was 16.5 g kgy1 dry matter, or 0.76 g MJy1 digestible energy, both close to the lower range of the estimated requirements for lysine

Ž .

in salmon Berge et al., 1998 . A consequence of the high protein level used in practical salmon diets, thus, is that the lysine requirement for growth was nearly or fully met in salmon fed the diet with 50% of CP from gluten. Some of absorbed lysine probably is deaminated and metabolised, and it is possible that supplementation with lysine would be needed in order to facilitate rapid growth and efficient feed conversion in salmon fed a diet with as much as 50% of CP from WG.

Based on the same assumptions used for indicating lysine requirement based on composition of growth and digestible amino acid intake, the requirements for all other essential amino acids seemed to have been met with solid margins in all diets. In the diet

Ž . Ž .

with 50% CP from WG, histidine 60% excess and threonine 70% excess appeared to be the 2nd and 3rd limiting amino acids. The first limiting essential amino acid in the

Ž .

FM control diet appeared to be histidine 50% in excess .

The negative apparent absorption estimates for Ca, Mg, and Sr illustrates that those elements were taken up from the water and excreted in the gut. There are previous examples of negative absorption values for Ca, even when the fish are reared in

Ž .

freshwater Storebakken, 1985 . In spite of this negative absorption estimate, salmon is

Ž .

able to maintain homeostatic regulation of Ca Storebakken et al., 1998 . Ca is efficiently taken up from the water, and water-borne Ca, from drinking or excretion by the intestine, is the source of the negative absorption. Uptake of Sr closely resembles

Ž .

that of Ca Yamada and Mulligan, 1987 . Salmonids are also able to take up Mg

˚

ŽShearer and Asgard, 1992 and Zn Spry et al., 1988 from the water. In contrast to

˚

. Ž . what has been found for diets with 75% of the protein from soy protein concentrate ŽStorebakken et al., 1998 , the diets with WG did not result in any drastic increase in the. faecal output of Na.

The P absorption at 35% in the diets was slightly lower than the 41% previously obtained with an extruded LT-94 FM control diet fed to Atlantic salmon in saltwater Ž_{Storebakken et al., 1998 . This previous diet contained 19.5 g P kg}. y1_{, and the sources}

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T. Storebakken et al.rAquaculture 184 2000 115–132 129

contained 15.5 g P kgy1, from 568 g FM and 13.5 g dicalcium phosphate kgy1. The difference most probably is rationalised by differences in availability of P from the FM

source, which has been documented in experiments with rainbow trout Riche and .

Brown, 1996 , and higher availability of P from the added dicalcium phosphate ŽNordrum et al., 1997 . The availability of P in the present experiment was equal to that.

Ž .

obtained by Hillestad et al. 1999 using a diet with a solvent-extracted LT-94 FM to salmon in saltwater. This diet contained 15.3 g kgy1_{, mainly originating from the FM,}

and the diet had no inorganic P added. The lack of reduced absorption of Ca, P or Mg in the salmon fed the diets with WG is logical, based on the low concentration of phytic acid present in feed ingredients and diets.

The absorption of Zn in the FM control diet was lower than found previously in

Ž .

Atlantic salmon Storebakken et al., 1998 , while the values for the diets with gluten were slightly lower than those obtained with phytase-treated soy protein concentrate, and more than twice the values obtained with 75% of the protein from untreated soy concentrate. A direct comparison is, however, limited by the concentration of Zn being almost 100 mg kgy1 lower in the present than in the previous study. Sugiura et al. Ž_{1997 also found that Zn from WG was highly absorbable in rainbow trout.}.

The present study does not give any explanation for the improved absorption of Zn when WG is included in the diets. Absorption of Zn is mediated by low-molecular-weight

Ž .

substances peptides or amino acids originated from protein digestion or

feedback-regu-Ž .

lated pancreatic secretions in rats Evans et al., 1975; Wapnir and Stiel, 1986 and Ž

further regulated by metallothionin in the epithelial cells in lower vertebrates Hogstrand

. Ž

and Haux, 1991 . Zn is mainly excreted through the gills in rainbow trout Hardy et al., .

1987 . Increased absorption and a homeostatic plateau for intestinal uptake appear to have been reached at the lowest level of gluten.

The inclusion of WG affected the physical quality of the pellets. All pellets absorbed fat well, but the density of the diets increased with increasing inclusion of WG. Adjustments were made in the extrusion parameters to get a good physical quality, and it may be that the deviations from the general pattern in ADC of macronutrients was the result of a modification in the extrusion process rather than depending on the nutritional qualities of the feed ingredients.

In this study, the difference in intestinal morphology between groups fed a FM diet and a diet with extracted SBM was clearly established, although the SBM-induced

Ž .

changes were less severe than previously observed by Baeverfjord and Krogdahl 1996 .

Ž .

There was, however, a difference between the soybean inclusion levels in this 15% and

Ž .

the previous 40% study. For the diet with WG, the small changes observed in one of the 15 individuals were non-specific and moderate, and were considered to be of little relevance to the question in issue.

5. Conclusion

WG seems to be a useful feed ingredient in grower diets for Atlantic salmon. WG supported rapid growth and was highly digestible, did not reduce digestibility of fat or

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energy or availability of essential elements, but a slight reduction of digestibility of starch was seen. The results also indicate that WG at a level up to 35% of dietary protein may be used with no adverse effect on the mucosa of the distal intestine.

Acknowledgements

The experiments were supported by grants from Amylum, Aalst, Belgium and BioMar, Myre, Norway.

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116, 2171–2179.

Wilson, R.P., Cowey, C.B., 1985. Amino acid composition of whole body tissue of rainbow trout and Atlantic salmon. Aquaculture 48, 373–376.

Yamada, S.B., Mulligan, T.J., 1987. Marking unfed salmonid fry with dissolved strontium. Can. J. Fish. Aquat. Sci. 44, 1502–1506.

Yamamoto, T., Akiyama, T., 1995. Effect of carboxymethylcellulose, alpha-starch, and wheat gluten incorpo-rated in diets as binders on growth, feed efficiency, and digestive enzyme activity of Japanese flounder. Fish. Sci. 61, 309–313.

(1)

Ž .

be ascribed to the a-amylase inhibitor found in wheat Hofer and Sturmbauer, 1985 .

This inhibitor is a protein, which may be concentrated in the protein fraction when producing the gluten.

The ADC of protein in the FM control diet agrees with previous findings with LT-94

FM in extruded high-energy diets fed to Atlantic salmon in saltwater Johnsen et al.,

1993; Aksnes, 1995 . The AAC of amino acids was generally 2–3% units higher than the results reported for a cold-pelleted LT-70 FM-based diet with approximately 180 g

y1 _Ž _.

Ž Ž 2_..

was 50% of the protein from WG, and that 12% 100= 1yr of the variation in

ADC of protein was due to other factors than the diet, or interactions among dietary components. The estimated ADC of CP, using the regression in Table 7, for a diet with WG as the only source of protein was 100%, in keeping with the value of 99% obtained

Ž .

by Pfeffer et al. 1995 in rainbow trout. The ADC of protein for the diet without gluten was 88.8%.

ŽBatterham, 1994 . It is probable that the tendency of lowered AAC of lysine with.

increasing dietary gluten level is explained by the endogenous lysine excretion repre-senting a relatively higher proportion of faecal amino acids with increasing dietary gluten levels.

Availability of cysteine in feed ingredients for salmonids may be low Skrede et al.,

1981, 1998 due to the negative effects of heat when the feed ingredient is dried, such as

Ž .

denaturation, oxidation and cross-linkage Opstvedt et al., 1984 . Furthermore,

sulphur-Ž .

There is limited information available on the amino acid requirement of Atlantic salmon reared in saltwater, but the requirement of lysine has recently been established to

16–18 g kgy1 dietary dry matter or 0.79–0.89 g MJy1 digestible energy in slowly

Ž .

growing salmon with an initial weight of 0.4 kg Berge et al., 1998 . These workers observed mortality and poor growth in the beginning of the study, and SGR was only

0.69 at 88C in the best group at the end of the experiment. When compared to

Ž .

whole-body amino acid profiles in Atlantic salmon Wilson and Cowey, 1985 , lysine is

Ž .

(2)

T. Storebakken et al.rAquaculture 184 2000 115–132

128

The optimal supplementation of lysine and eventually other essential amino acids must be resolved in future experiments with Atlantic salmon.

Ž .

Atlantic salmon constitutes of 180 g CP Shearer et al., 1994 . Whole-body protein of

Ž .

Atlantic salmon contains 9.28 g lysiner100 g amino acids Wilson and Cowey, 1985 . If

Ž .

the amino acid-N accounts for 92% of CP Storebakken, unpublished , the net lysine

requirement for growth, thus, should be approximately 15.4 g kgy1 _{gain. The lysine}

content of the diet with 50% protein from WG was 3.6 gr100 g CP, the dietary CP

concentration was 458 g kgy1_{, and AAC of lysine was 95.8. If assuming a feed}

conversion ratio of 1 kg dry feed intake per kilogram gain, the digestible lysine intake

would be 15.8 g kgy1

gain. The gross lysine content in the diet with 50% of CP from

WG, not corrected for digestibility, was 16.5 g kgy1 dry matter, or 0.76 g MJy1

digestible energy, both close to the lower range of the estimated requirements for lysine

Ž .

Ž . Ž .

with 50% CP from WG, histidine 60% excess and threonine 70% excess appeared to be the 2nd and 3rd limiting amino acids. The first limiting essential amino acid in the

Ž .

FM control diet appeared to be histidine 50% in excess .

Ž .

freshwater Storebakken, 1985 . In spite of this negative absorption estimate, salmon is

Ž .

able to maintain homeostatic regulation of Ca Storebakken et al., 1998 . Ca is

efficiently taken up from the water, and water-borne Ca, from drinking or excretion by the intestine, is the source of the negative absorption. Uptake of Sr closely resembles

Ž .

that of Ca Yamada and Mulligan, 1987 . Salmonids are also able to take up Mg

˚

ŽShearer and Asgard, 1992 and Zn Spry et al., 1988 from the water. In contrast to

˚

. Ž .

what has been found for diets with 75% of the protein from soy protein concentrate

ŽStorebakken et al., 1998 , the diets with WG did not result in any drastic increase in the.

faecal output of Na.

The P absorption at 35% in the diets was slightly lower than the 41% previously obtained with an extruded LT-94 FM control diet fed to Atlantic salmon in saltwater

Ž_{Storebakken et al., 1998 . This previous diet contained 19.5 g P kg}. y1_{, and the sources}

(3)

contained 15.5 g P kgy1, from 568 g FM and 13.5 g dicalcium phosphate kgy1. The difference most probably is rationalised by differences in availability of P from the FM

source, which has been documented in experiments with rainbow trout Riche and

Brown, 1996 , and higher availability of P from the added dicalcium phosphate

ŽNordrum et al., 1997 . The availability of P in the present experiment was equal to that.

Ž .

obtained by Hillestad et al. 1999 using a diet with a solvent-extracted LT-94 FM to salmon in saltwater. This diet contained 15.3 g kgy1_{, mainly originating from the FM,} and the diet had no inorganic P added. The lack of reduced absorption of Ca, P or Mg in the salmon fed the diets with WG is logical, based on the low concentration of phytic acid present in feed ingredients and diets.

The absorption of Zn in the FM control diet was lower than found previously in

Ž .

almost 100 mg kgy1 lower in the present than in the previous study. Sugiura et al.

Ž_{1997 also found that Zn from WG was highly absorbable in rainbow trout.}.

The present study does not give any explanation for the improved absorption of Zn when WG is included in the diets. Absorption of Zn is mediated by low-molecular-weight

Ž .

substances peptides or amino acids originated from protein digestion or

feedback-regu-Ž .

lated pancreatic secretions in rats Evans et al., 1975; Wapnir and Stiel, 1986 and

further regulated by metallothionin in the epithelial cells in lower vertebrates Hogstrand

. Ž

and Haux, 1991 . Zn is mainly excreted through the gills in rainbow trout Hardy et al.,

1987 . Increased absorption and a homeostatic plateau for intestinal uptake appear to have been reached at the lowest level of gluten.

In this study, the difference in intestinal morphology between groups fed a FM diet and a diet with extracted SBM was clearly established, although the SBM-induced

Ž .

changes were less severe than previously observed by Baeverfjord and Krogdahl 1996 .

Ž .

There was, however, a difference between the soybean inclusion levels in this 15% and

Ž .

5. Conclusion

WG seems to be a useful feed ingredient in grower diets for Atlantic salmon. WG supported rapid growth and was highly digestible, did not reduce digestibility of fat or

(4)

T. Storebakken et al.rAquaculture 184 2000 115–132

130

Acknowledgements

The experiments were supported by grants from Amylum, Aalst, Belgium and BioMar, Myre, Norway.

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Directory UMM :Data Elmu:jurnal:A:Aquaculture:Vol184.Issue1-2.Apr2000:

Digestibility of macronutrients, energy and amino

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