Nutritional and Metabolic Characteristics of Brassica carinata Co

Jun 26, 2017 - The objectives of this study were to investigate the nutritive value of carinata presscake and meal for dairy cows in terms of (1) nutr...
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Nutritional and Metabolic Characteristics of Brassica Carinata Coproducts from Biofuel Processing in Dairy Cows Yajing Ban, Nazir Ahmad Khan, and Peiqiang Yu J. Agric. Food Chem., Just Accepted Manuscript • DOI: 10.1021/acs.jafc.7b02330 • Publication Date (Web): 26 Jun 2017 Downloaded from http://pubs.acs.org on June 27, 2017

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Journal of Agricultural and Food Chemistry is published by the American Chemical Society. 1155 Sixteenth Street N.W., Washington, DC 20036 Published by American Chemical Society. Copyright © American Chemical Society. However, no copyright claim is made to original U.S. Government works, or works produced by employees of any Commonwealth realm Crown government in the course of their duties.

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Nutritional and Metabolic Characteristics of Brassica Carinata Coproducts from Biofuel

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Processing in Dairy Cows

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Yajing Ban1, Nazir Ahmad Khan1,2,*, and Peiqiang Yu1,**

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University of Saskatchewan, Saskatoon, SK, S7N 5A8, Canada

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2,

Department of Animal and Poultry Science, College of Agriculture and Bioresources,

Department of Animal Nutrition, The University of Agriculture Peshawar, 25130, KP, Pakistan

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*

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Tel: +1 306 966 4132; Fax: +1 306 966 4150

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E-mail: [email protected]

Corresponding author at manuscript submission stage:

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** Corresponding author

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Peiqiang Yu, Ph.D.

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Professor and Ministry of Agriculture Strategic Research Chair

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College of Agriculture and Bioresources, University of Saskatchewan

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6D10 Agriculture Building, 51 Campus Drive, Saskatoon

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Canada, S7N 5A8

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Tel:

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E-mail:[email protected]

+1 306 966 4132; Fax:

+1 306 966 4150

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ABSTRACT: The increased utilization of Brassica carinata in the biofuel industry in Canada

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has resulted in the large scale production of coproducts that can be potentially exploited as

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alternative protein ingredient in dairy ration. The objectives of this study were to investigate the

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nutritive value of carinata presscake and meal for dairy cows in terms of (1) nutrient and anti-

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nutrient composition; (2) rumen degradation kinetics of organic matter (OM), crude protein (CP)

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and neutral detergent fiber; (3) hourly effective degradation ratio and potential N to energy

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synchronization; (4) intestinal digestion of rumen undegraded protein (RUP); and (5) total

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metabolizable protein (MP) supply to the small intestine. Samples (n=3) of carinata meal,

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carinata presscake and canola meal (as reference feed), collected from three consecutive batches,

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were evaluated. Compared to canola meal, carinata presscake and meal had greater (P < 0.05)

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contents of CP (39.7% vs. 48.5%, 53.5% dry matter (DM)), with a high proportion of soluble

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crude protein (24.0% vs. 53.0%, 72.6% CP), resulting in their extensive degradation (59.2% vs.

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76.3, 89.3% CP) in the rumen. As a result, carinata presscake and meal supplied smaller (P
99%),

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sodium tungstate (Na2WO4·2H2O; purity 100%), boric acid (purity > 99.5%), sulphuric acid

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(purity 70%), ethyl ether (purity 99%), sodium borate (Na2B4O7·10H2O; purity 99.5%),

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hydrochloric acid (purity 38%), acetone (purity ≥ 99.5%), potassium phosphate (purity > 99%),

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sodium sulfite anhydrous (purity 98.4%), kjeltabs (0.15 g CuSO4 and 5.0 g K2SO4; N < 0.005%)

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and potassium hydroxide (purity 85%) were purchased from Fisher Scientific (Ottawa, ON,

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Canada). The enzymes, pepsin and pancreatin, were purchased from Sigma-Aldrich Co. (St.

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Louis, MO, USA). The Neutral detergent fiber solution containing 30 g sodium dodecyl

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sulphate, 18.61 g ethylenediamine-tetraacetic acid disodium salt, 6.81 g sodium borate, 4.56 g

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anhydrous sodium phosphate diabasic,10 ml triethylene glycol and heat-stable α-amylase (17400

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liquefon units/ml), and acid detergent fiber (ADF) solution containing 20 g cetyl trimethyl

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ammonium bromide in 1 N sulphuric acid were purchased from Ankom Technology (Macedon,

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NY, USA).

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Carinata Processing

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In this study carinata meal and carinata presscake were evaluated in comparison with

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commercial canola meal as a reference feed. The cold-pressing of carinata seeds from three

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consecutive batches was carried out in POS Bio-Sciences (Saskatoon, SK) according to the

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approved method of Agrisoma Biosciences Inc. (Saskatoon, SK). From each batch three samples

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of carinata presscake were collected, and no added heating was used during the cold-pressing.

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Similarly, three batches of carinata meal were produced from seeds received in three consecutive

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batches at Agrisoma Biosciences Inc. (Saskatoon, SK). For the meal production, mixed-colored

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carinata seeds (yellow and brown) were weighed, transferred into a 4 L plastic pail, and sprayed

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with water to maintain moisture content of 2.5%. After thorough mixing, the tempered seeds

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were flaked using the lab flaking mill and heated in beakers in a microwave oven (DMW904W,

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Findlay, OH, US), operated at a power of 900 W (1.33 W/g) and an irradiation frequency of

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2450 MHz for 2.5 min. Then all beakers were covered and transferred to a conventional oven for

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20 minutes at 95±3 °C. Later, the cooked flakes were pressed through a 5/16 die plate using the

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Gusta Laboratory Press (Gusta Manufacturing, Winnipeg, MB), and extracted oil during the

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crude-pressed process was collected. The laboratory Soxhlet extractor was then used to extract

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the residual oil using 6 liters of fresh hexane for 4 h. Lastly, the carinata meal was placed in a

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fume hood for a minimum of 36 h for air desolventizing. Three samples were collected from

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each batch of carinata meal. Samples from three batches of canola meal (n=3 from each batch)

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were provided by Federated Cooperatives Limited (Saskatoon, SK) as a reference.

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Chemical Analysis

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For chemical analysis the meals and presscake samples were ground (Retsch ZM-1, Brinkmann

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Instruments Ltd., Mississauga, ON, Canada) through a 1-mm screen, whereas for in situ

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incubation studies the samples were ground through a 2-mm screen. The contents of dry matter

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(DM; method 930.15), ash (method 942.05), crude fat (Cfat; method 954.02) and CP (method

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984.13) were analyzed according to the official methods of AOAC.14 The neutral detergent fiber

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(NDF), acid detergent fiber (ADF) and acid detergent lignin (ADL) were analyzed according to

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the method of Van Soest et al.15 using the ANKOM A200 filter bag technique (ANKOM

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Technology Corp., Fairport, NY, US). Neutral detergent insoluble CP (NDICP) and acid

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detergent insoluble CP (ADICP) were analyzed according to Licitra et al.16 using the NDF and

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ADF residues. For determination of soluble CP (SCP) content, all samples were incubated in a

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borate-phosphate buffer for three hour, and filtered through #54 Whatman filter papers.16 Non-

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protein nitrogen (NPN) was estimated as the difference between total CP and precipitated true

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protein in tungstic acid.16 Total carbohydrate fraction (CHO) was calculated according to NRC 17

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as: CHO = 100 - (Cfat + CP + Ash). The contents of glucosinolates were determined by POS

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Bio-Sciences laboratories (Saskatoon, SK) according to the official method of the Canadian

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Grain Commission.18 The condensed tannins were determined using the HCl−butanol

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procedure.19

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Animals, Diets and Rumen Incubation

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Four rumen-cannulated (10 cm internal diameter; Bar Diamond Inc., Parma, ID, USA) non-

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lactating Holstein dairy cows (body weight, 680 ± 10 kg), were used for the in situ incubation

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trial. The procedures involved in the animal experiment was approved by the University of

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Saskatchewan Animal Research Ethics Board (Animal Use Protocol No. 19910012), and

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conducted according to the guidelines of the Canadian Council of Animal Care. 20 Cows were

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individually fed twice daily at 0800 and 1600 h with a total mixed ration (TMR) in a tie-stall

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barn at the Rayner Dairy Research and Teaching Facility (University of Saskatchewan,

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Saskatoon, SK), and given free access to water. The TMR was formulated with 56.5% barley

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silage, 12.2% hay and 31.3% concentrate according to the NRC requirements.17 The standard

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method of in situ rumen incubation as described by Khan et al.21 was used to determine the

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rumen degradation kinetics of the experimental feeds. The dried residues were weighed and

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pooled for chemical analyses as per treatment and incubation time within each run. The residues

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were ground by a coffee grinder (PC770, Loblaws Inc., Toronto, ON) for 10 s, repeated once, to

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obtain approximately 1 mm particle size. The residues were analyzed the content of CP and NDF

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as described earlier.

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Rumen Degradation Kinetics

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The first order kinetics degradation model with lag time was used to estimate the degradation

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features of OM, CP and NDF.22-24 The data was processed using the non-linear (NLIN)

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procedure of SAS 9.3 (SAS Institute, Inc., Cary, NC, US) and iterative least-squares regression

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(Gauss-Newton method). R  = U + D × e     7

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Where R(t) is the residue (%) after t h incubation, U is undegradable fraction (%), D is

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degradable fraction (%), Kd is degradation rate (%/h), and T0 is lag time (h).

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The rumen undegraded fractions of OM (RUOM), CP (RUP) and NDF (RUNDF) were estimated using a passage rate (Kp) of 6%/h17, 24: RU% = U + D × 



  



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where RU is rumen undegradable fractions of OM, CP and NDF. The rumen degradable

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fractions were estimated as: % =  +  × 



 +  ! 

"

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where ED is effective degradability of OM (EDOM), CP (RDP) and NDF (EDNDF), and S is

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soluble fraction (%).

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The hourly nutrient degradation was estimated based on the equations from Sinclair et al.

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Based on this study, the equation of hourly effective degradation ratios of N to OM was

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published by Nuez-Ortín and Yu.26

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Hourly ED (g/kg DM) = S + [D × Kd / (Kp + Kd)] × [1 − e−t × (Kd + Kp)],

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Hourly ED N/OMt = (HEDNt − HEDNt−1) / (HEDOMt − HEDOMt−1),

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where N/OMt is the ratio of N to OM at time t (g N/kg OM), HEDNt is effective

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degradability of N at time t (g/kg DM), HEDNt−1 is effective degradability of N at 1 h before

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time t (g/kg DM), HEDOMt is effective degradability of OM at time t (g/kg DM), and

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HEDOMt−1 is effective degradability of OM at 1 h before time t (g/kg DM).The optimum N to

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OM degradation ratio was considered to be 25 g N/kg OM to realize the most efficient microbial

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crude protein (MCP) synthesis. 26

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Intestinal Digestibility of Protein

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Digestibility of RUP in the small intestine was determined by three-step in vitro procedure of

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Calsamiglia and Stern.27 Briefly, 15 g of residues from 12 h in situ rumen incubation were

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weighed and mixed with 10 mL of 0.1 mol/L HCl solution (pH = 1.9) containing pepsin, and

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then incubated at 38 ºC for 1 h in a shaking water bath. After incubation, the solution was

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neutralized by the addition of 0.5 mL of 1 mol/L NaOH (pH = 7.8) and 13.5 mL of phosphate-

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buffer containing pancreatin. The mixture was subsequently vortexed, and incubated for 24 h at

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38 ºC in the shaking water bath, and vortexed every 8 h. At the end of 24 h incubation, 3 mL

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trichloroacetic acid (TCA) solution was added to the mixture in order to stop enzymatic activity

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and precipitate the undigested protein. Samples were then centrifuged for 15 min at 5000 rpm,

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and 5 ml of supernatant was analyzed for soluble N by the Kjeldahl method. The soluble protein

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was regarded as the digestible protein in the small intestine.

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Predicted Truly Absorbed Protein Supply and Feed Milk Value

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The metaboliable protein (MP) supplied to dairy cows was calculated as the sum of (1) truly

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absorbable rumen synthesized MCP in the small intestine (AMCP); (2) truly absorbable rumen-

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undegradable feed protein reaching the small intestine (ARUP); and (3) truly absorbable

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endogenous protein reaching the small intestine (AECP).17 The detailed description of the

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concepts, equations and formulas for estimation of AMCP, ARUP and AECP are given by Peng

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et al.28 The MP absorbed from the small intestine is partly used for milk production. The feed

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milk value (FMV) is the term used for estimating the efficiency of feed MP for milk

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production.29 The transfer efficiency of MP to milk is 0.67, and 1 kg milk contains

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approximately 33 g milk protein 17,29 Therefore, FMV can be estimated as:

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FMV = 0.67 ×

MPg/kg DM 33

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Statistical Analysis

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Data on the chemical profile, rumen degradation kinetics of nutrients, hourly effective

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degradation ratios, intestinal digestion of protein, MP supply to the small intestine and FMV of

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the carinata coproducts and canola meal were analyzed using the MIXED procedure of SAS 9.3

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(SAS Institute, Inc., Cary, NC, US). The model used for the analyses of nutrients and anti-

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nutrients profile was

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Yij = µ + Fi + eij,

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where Yij is an observation of the dependent variable ij, µ is the population mean for the

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variable, Fi is the effect of different coproducts as a fixed effect (different sources were treated as

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the replication), and eij is the random error associated with the observation ij. For the analysis of

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rumen degradation kinetics, hourly effective degradation ratios, intestinal digestion of protein

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and MP supply the following model was used,

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Yijk = µ + Fi + Sj + eijk,

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where Yijk is an observation of the dependent variable ijk, µ is the population mean for the

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variable, Fi is the effect of different coproducts as a fixed effect, Sj is the effect of different

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batches in the in situ trial as a random effect, and eijk is the random error associated with the

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observation ij. For all statistical analyses, significance was declared at P < 0.05, and trends at

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0.05 ≤ P ≤ 0.10.

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Results and discussion

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Chemical Profiles and Anti-nutritional Compounds of Carinata Coproducts 10

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The results of chemical profile, protein and carbohydrate sub-fraction compositions of B.

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carinata coproducts, meal and presscake, in comparison with canola meal are presented in Table

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1. There were no significant differences among carinata coproducts and canola meal for the

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contents of ash. Carinata presscake had a higher (P < 0.05) content (2.5% DM) of Cfat than

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carinata meal (0.3% DM). The presscake was produced after partial extraction of oil from

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carinata seeds by cold-pressing; whereas, from meal the residual oil was further removed by

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solvent-extraction process. This explains the higher content of Cfat in carinata presscake as

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compared to carinata meal. In agreement with our findings Theodoridou and Yu30 reported a

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higher content of Cfat in canola presscake as compared to canola meal. Notably, both coproducts

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of carinata had higher (P < 0.05) contents of CP than canola meal. A further comparison between

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the carinata coproducts revealed that the presscake had numerically lower content (48.5% vs.

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53.5% DM) of CP than the meal. This result is in accordance with the literature,31,32 and a

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plausible explanation is that the cold-press system of oil extraction results in lower oil recovery

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from the presscake compared to the solvent extraction, and as a result the presscake contains a

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higher content of oil and lower content of CP as compared to the meal.

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The CP subfraction composition revealed that compared to canola meal the SCP contents were

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more than two times higher (P < 0.05) in the carinata coproducts. The SCP is expected to

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degrade rapidly in the rumen, and may not be optimally utilized for MCP synthesis, that can

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lower the overall supply of MP to the dairy cow.33,34 Between the carinata coproducts, the

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presscake had higher (P < 0.05) SCP content (72.6% vs. 53.0% DM) than carinata meal. Mustafa

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et al.35 reported that compared to presscake the SCP content in solvent extracted meal is reduced

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by more than half after toasting, which they attributed to both high moisture and high heating,

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that prevailed during the solvent extraction process. Heating during solvent extraction can alter

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protein structures such as by the uncoiling of the pleated structures or by denaturing the protein

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molecules.34,36 The subsequent liberation of hydrophobic amino acids, racemization, and

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establishment of cross-linkages among the uncoiled peptides and between amino acids and

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reducing sugars can all reduce CP solubility. The contents of NDIP and ADIP were lower (P
0.10) in the Kd

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among the three coproducts. The greater (P < 0.05) D-fraction and smaller (P < 0.05) U-fraction

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was recorded for carinata presscake, while carinata and canola meal did not differ (P > 0.05) in

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these degradation characteristics. Among the three coproducts, carinata presscake had greater (P

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< 0.05) EDNDF (47.1% NDF), while carinata and canola meal did not differ (P > 0.05) in

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EDNDF. Our findings are consistent with literature values of McKinnon and Walker.31 However,

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another study indicated that presscake processing may not change rumen degradability of NDF.29

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Hourly Effective Rumen Degradation Ratios of Carinata Coproducts

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The optimal effective degradation ratio between N and energy supply to the rumen of dairy cows

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is 25g N/kg OM for achieving maximum MCP synthesis and minimum N loss to environment.

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lower degradation ratio indicates a shortage of N supply for MCP synthesis. Figure 1 shows the

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hourly effective N/OM degradation ratios of the three coproducts at different incubation times.

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The hourly effective N/OM degradation ratios of the two carinata coproducts were relatively

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higher than that of canola meal at the beginning (0 h) of incubation, with only the ratio of canola

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meal being less than the optimum (25g N/kg OM). Subsequently, a dramatic increase in the

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N/OM effective degradability ratio was observed between 0 to 2 h for the three coproducts, and a

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greater change (21 to 92g N/kg OM) was observed for canola meal. In the first 2 h the changes in

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N/OM degradation ratios of carinata meal and presscake were more or less similar and both

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attained a similar ratio (120g N/kg OM) after 2 h of incubation. However, compared to carinata

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meal, the N/OM degradation ratios of carinata presscake decreased rapidly from 120 g N/kg OM

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at 2 h to 14 g N/kg OM at 24 h, which is less than the optimum ratio (