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Protein Concentrate Production from Thin Stillage Kornsulee Ratanapariyanuch, Youn Young Shim, Shahram Emami, and Martin J.T. Reaney J. Agric. Food Chem., Just Accepted Manuscript • DOI: 10.1021/acs.jafc.6b03816 • Publication Date (Web): 28 Nov 2016 Downloaded from http://pubs.acs.org on November 29, 2016

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Journal of Agricultural and Food Chemistry

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Protein Concentrate Production from Thin Stillage

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Kornsulee Ratanapariyanuch,† Youn Young Shim,*,†,‡,§ Shahram Emami,† Martin J. T.

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Reaney*,†,‡,§

5 6

†

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Saskatchewan S7N 5A8, Canada

8

‡

9

§

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Department of Plant Sciences, University of Saskatchewan, 51 Campus Drive, Saskatoon, Prairie Tide Chemicals Inc., 102 Melville Street, Saskatoon, Saskatchewan S7J 0R1, Canada

Guangdong Saskatchewan Oilseed Joint Laboratory, Department of Food Science and Engineering,

Jinan University, 601 Huangpu Avenue West, Guangzhou, Guangdong 510632, China

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AUTHOR INFORMATION

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E-mail address: [email protected] (KR), [email protected] (YYS),

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[email protected] (SE), [email protected] (MJTR) 1 ACS Paragon Plus Environment


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ABSTRACT

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Two-stage fermentation (TSF) of saccharified wheat with a consortium of endemic lactobacilli produced

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CO2 and induced colloid separation of fermented solution to produce a protein concentrate (PC).

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Protein-rich slurry (50%, db) was obtained by decanting solution or skimming floating material during

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or after the TSF. Washing and drying processes were explored to improve protein content, extend

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storage life of slurry and yield converted stillage for compound recovery. Centrifuging and washing

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slurry afforded a PC and clarified solution. PC protein content increased to 60% (w/w, db). The PC was

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dried in a spray dryer or drum dryer or tray dryer. Dried PC water activity ranged 0.23−0.30. The dried

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PC lysine content was low but lysine availability (95%) was excellent. Liquid from TSF and washing

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was readily microfiltered. Mass balance of protein, glycerol, 1,3-propanediol, lactic acid, acetic acid,

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and glycerylphosphorylcholine from combined TSF, washing, and filtration were 66, 76, 72, 77, 74, and

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84%, respectively.

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Keywords: Clarification; Compound recovery; Protein concentrate; Thin stillage; Two-stage

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fermentation

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2 ACS Paragon Plus Environment

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INTRODUCTION

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The production of ethanol by yeast fermentation followed by distillation produces a dilute aqueous

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stream (thin stillage, TS) of inorganic salts, organic solutes, and suspended particles.1−3 Water in TS can

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be evaporated to concentrate TS to distillers’ solubles (DS). DS, an inexpensive material, is sold or

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mixed with wet grain to produce dried distillers’ grain with solubles (DDGS), an animal feed

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ingredient.4 For example, the price of corn-based DS varied from 7.4 to 51.8 CAD $/tonne depending on

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ethanol manufacturer.5 In addition, concentration of TS to DS by evaporation is not energy efficient, as

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this process consumes 40−45% of all thermal energy and 30−40% of all electrical energy utilized in dry

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grind ethanol production facilities.6 Therefore, research has been conducted to utilize corn-based TS and

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wheat-based TS (W-TS) as animal feed.7,8 One ethanol plant utilizes W-TS as a water source for

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animals (e.g. Pound-Maker Agventures, Lanigan, SK, Canada). The use of energy for water evaporation

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is greatly reduced with this practise when compared to more conventional production practises.

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As previously stated, corn-based DS can be utilized as an animal feed ingredient. Corn-based DS

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contains essential amino acids and 18−23% protein (db). Lysine varied from 0.8−1.3% of protein.9

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Corn-based DS lysine content is lower than in soy protein concentrates (4.23%).10,11 Clarification of W-

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TS was described in Ratanapariyanuch et al.12 It was discovered that slurry obtained from two-stage

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fermentation (TSF) contained high protein (approximately 50% w/w, db) with lactobacilli contributing

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more than 93% of endemic flora present in the slurry. These observations indicate that the slurry

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fraction had higher protein concentration (db) than W-TS and that TSF could be used to produce a W-

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TS protein concentrate (PC)12 which could be utilized as protein source for animal feed. Lactobacilli are

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considered non-toxic bacteria and could have probiotic effects, as described in Pedersen et al.,13 when

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the slurry is utilized as an animal feed protein source. However, moisture present in slurry could

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increase shipping costs and lead to spoilage. Furthermore, valuable solutes dissolved in liquids

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surrounding the slurry might be recovered from the PC before drying.



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Slurry washing involves centrifugation in a decanter then mixing slurry with water followed by

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decanting again. Centrifugation accelerates the particle sedimentation rate14 and centripetal force could

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generate pressure that removes liquid surrounding particles and generates sediment with increased

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density. Water added to sediments could also extract soluble compounds from liquid surrounding

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centrifuged sediments and enable their recovery. These methods were also utilized to recover bacteria

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for DNA extraction as employed by Steffan et al.15

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Liquid from TSF and from washing could be combined for compound extraction and compound

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recovery. According to Ratanapariyanuch16 and Reaney et al.,17 the concentration of 1,3-propanediol

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(1,3-PD) and acetic acid in fermentation medium increased, while the concentration of glycerol and

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lactic acid decreased. If 1,3-PD and acetic acid were extracted from TSF liquid fractions and slurry

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water wash, the water after compound extraction would be less acidic (less acetic acid). Acetic and

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lactic acids cause yeast stress that may lead to cell death.18 Therefore, liquid that is depleted in lactic and

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acetic acid produced by TSF and compound extraction might provide better recycle water for ethanol

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production (backset). After washing, slurries from TSF contained 86−87% moisture. Therefore, drying

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processes are necessary to prevent spoilage and reduce shipping costs. In addition, qualities of dried PC

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should be determined prior to use as a protein source in animal feed.

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The objectives of this study were 1) to study washing processes at both a laboratory and pilot-scale

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with the goals of increasing protein content and recovering organic solutes present in W-TS including

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1,3-PD, acetic acid, and alpha-glycerylphosphorylcholine (GPC) and 2) to study the possibility of using

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three different drying methods: spray drying, drum drying, and tray drying for drying PC to determine

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the qualities of dried PC including crude protein and moisture contents, water activity (aw), crude fat,

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ash, fiber, acid detergent fiber, neutral detergent fiber, amino acid profile, lysine availability, and

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mineral content.


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MATERIALS AND METHODS

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Materials and Equipment.

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TSF of W-TS with endemic microorganisms was performed at 25 °C to produce slurry I and liquid I

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for further processes. Slurry I samples from small-scale fermentation at 25 °C replicates 1 and 212 were

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utilized to study slurry washing (30 mL; hereafter called laboratory processing). Slurry I samples

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produced by decanting twelve fermenters (25 L each) were employed to study slurry washing at a pilot-

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scale (230 kg/replicate)12 using continuous processing equipment (POS Biosciences, Saskatoon, SK,

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Canada). A solid-bowl decanter (CA225-010, Westfalia, GEA Mechanical Equipment US Inc.,

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Northvale, NJ, USA) and a disc stack desludger centrifuge (SA7-06-576, Westfalia, GEA Mechanical

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Equipment US Inc., Northvale, NJ, USA) were utilized for washing slurry I (300 L fermentation

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volume) to produce PC. In addition, a digital scale (MS 5060S, Ishida Digital Scale, Ishida Co. Ltd.,

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Sakyo-ku, Kyoto, Japan) was used to record wash fraction masses.

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A crossflow filter (Sartocon2 plus 1−10 cassette system, Sartorius AG, Gӧettingen, Germany), with 5

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spaced cassettes (0.6 m2, wide spacer) and microfiltration membranes (pore size 0.2 µm) and a standard

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industrial three phase motor and pump (CM3558T, Baldor electric Co., Fort Smith, AR, USA) were

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utilized to filter TSF liquids and liquid fractions from washing slurry in pilot-scale replicate 1. Spray

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drying (B-290, BÜCHI Labortechnik AG, Flawil, Switzerland), drum drying (Buflovak, Buflovak

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equipment division of Blaw-knox Co., Buffalo, NY, USA), and forced-air drying (1390 FM, Sheldon

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Manufacturing Inc., Cornelius, OR, USA) were employed to produce dried PC.

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Analytical Methods.

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Protein and Moisture Contents. Protein contents of slurry and wash fractions of both laboratory

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scale and pilot-scale processes were determined using the Kjeldahl method (AOAC 981.10)19 as

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modified from Ratanapariyanuch et al.3 Nitrogen content present in non-protein compounds (GPC and

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betaine) was subtracted from total nitrogen content prior to calculating protein content.3 Nitrogen

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content was used to estimate protein content by multiplication by 5.7.20 Moisture content was 5 ACS Paragon Plus Environment


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determined according to AOAC 950.4619 by oven drying samples to a constant mass at 100 ± 2 °C for

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16−18 h as modified from Ratanapariyanuch.2

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Double Pulse Field Gradient Spin Echo-NMR Spectrometry. Double pulse field gradient spin echo

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proton (DPFGSE-NMR) was conducted to quantify and characterize organic solutes in solutions

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according to a modified method based on Ratanapariyanuch et al.3 Samples were centrifuged

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(Spectrafuge™ 24D, Labnet International Inc., Edison, NJ, USA) at 9200g for 10 min prior to analysis.

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After centrifugation, supernatant samples were filtered through syringe filters (25 mm syringe filter with

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0.45 µm PTFE membrane, VWR International, West Chester, PA, USA). Proton DPFGSE-NMR

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spectra were recorded using a Bruker Avance 500 MHz NMR spectrometer (Bruker BioSpin,

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Rheinstetten, Germany) with 16 scans per spectrum. NMR data collection and analysis were conducted

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with TopSpin™ 3.2 software (Bruker BioSpin GmbH, Billerica, MA, USA). Deuterium oxide

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(Cambridge Isotope Laboratories Inc., Andover, MA, USA) and dimethylformamide (EMD Chemicals

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Inc., Gibbstown, NJ, USA) were used as solvent and internal standard, respectively.

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Processing for PC Production.

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Laboratory Processing. Laboratory processing (Figure 1) of TSF product (slurry I, 30 mL) was

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conducted with a modified method of Steffan et al.15 The slurry I was centrifuged using a benchtop

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centrifuge (Allegra X-22R, Beckman Coulter Canada Inc., Mississauga, ON, Canada) at 4 °C and 800g

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for 10 min to obtain solid I and liquid II fractions. Solid I was mixed with reverse osmosis (RO) water at

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a ratio of 1: 1 (v:v) and stirred for 30 min. The solution was then centrifuged at 800g for 10 min at 4 °C.

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Supernatant was decanted, hereafter called liquid III. The solid pellet remaining after centrifugation,

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hereafter called solid II, was washed by suspending in water, centrifugation then decanting. The pellet

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was washed twice using this procedure. Supernatants recovered after the 2nd and 3rd washes were called

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liquid IV and liquid V. The pellet after the final wash is called solid IV and it is also referred to as PC.

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A second replicate was conducted and the results are available in supplemental data. Nitrogen and

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moisture contents of liquid II−liquid V and solid IV were analyzed in duplicate. In addition, the 6 ACS Paragon Plus Environment

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concentrations of organic solutes in liquid II−liquid V were analyzed using DPFGSE-NMR to determine

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organic solutes in single analysis of each replicate.

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Pilot-scale Processing. Based on protein content and the concentration of organic solutes from

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washing slurry I observed in laboratory scale processing, it was concluded that 2 washes were sufficient

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to increase protein content to 60% (w/w, db) and recover valuable compounds (1,3-PD, acetic acid, and

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GPC). Therefore, TSF product (slurry I, 230 kg) was washed only 2 times during pilot-scale processing

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(Figure 2). Pilot-scale washing was conducted at POS Bio-Sciences as shown in Figure 2. Slurry I

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sample was passed through a solid-bowl decanter (3322g) to remove particles and produce a fraction

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called solid I decanter. Liquid from the decanter was passed through a disc stack desludger centrifuge

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(6540g) to remove fine particles (producing solid I desludger). After decanting and desludging

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treatments the resulting supernatant fractions were called liquid I decanter and liquid I desludger,

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respectively. Solid I decanter and solid I desludger were then mixed with water at a ratio of 1: 1 (w: w)

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in a mixing tank for 30 min for the 1st wash. The mixture was then passed through the decanter and

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desludger as previously described. Solids and supernatant from the 1st wash were called solid II

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decanter, solid II desludger, liquid II decanter, and liquid II desludger, in that order. Solid II decanter

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and solid II desludger were mixed and washed for the 2nd time using the same procedure used for the 1st

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wash. Solid and supernatant from the 2nd wash were called solid III decanter, solid III desludger, liquid

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III decanter, and liquid III desludger. The mass of each wash fraction from washing process was

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recorded. Decanter and desludger feed rates for the 1st pass, 1st wash, and 2nd wash are presented in

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Table 1. The pilot-scale washing process was repeated in its entirety as a separate replicate (replicate 2).

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Samples of all washing fractions were collected for determination of protein and moisture contents in

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duplicate. The concentration of organic solutes in liquid fractions were determined using DPFGSE-

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NMR in single analysis. The replicates were produced from different stillage batches and the data were

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not directly compared statistically.



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Crossflow Filtration of Liquid Fractions. Liquid I, liquid I desludger, liquid II desludger, and liquid

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III desludger from TSF and washing from replications of TSF replicate 1 were filtered through a

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crossflow filter. The flow was provided with a standard industrial three phase motor and pump. The

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crossflow filter was operated at inlet, permeate and retentate pressures of 173−207, 49 and 69 kPa,

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respectively, as recommended by the manufacturer. The motor was operated at 1,784 rpm. The

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permeate and retentate masses and filtration times were recorded. Permeate flux was also determined.

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Permeate and retentate were analyzed for protein and moisture contents in duplicate and concentration

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of organic solutes was determined as a single analysis.

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Drying PC. Solids produced from washing slurry I (pilot scale) were dried using either a spray dryer,

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drum dryer, or tray dryer to produce dried PC. The wet PC feed supplied to the spray dryer or drum

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dryer was mixed with RO water at a ratio of 1: 1 (w: w) to aid flow. The inlet temperature for spray

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drying was adjusted to control the outlet temperature, while aspirator, % pump, and nozzle cleaning

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were set at 100, 20, and 4, respectively. While feeding PC solution the set point inlet temperature was

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adjusted to 149, 139, and 129 °C. The outlet temperatures were 90, 83, and 75 °C, respectively, under

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these operating conditions. During drum drying, drum rotation speed was adjusted to control drum

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surface temperature and contact time, while stream pressure was set at 221 kPa. The drum rotation

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speed was adjusted to 2 (78 s/revolution) and 1 (125 s/revolution). Samples were placed in Handi-foil

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aluminum steam table pans for tray drying. Wet PC was added at a depth of 1 cm. The oven temperature

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was 70 °C to obtain dried PC.

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Dried PC color was determined in duplicate using a Hunterlab Miniscan XE (Hunter, Reston VA,

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USA). Color was expressed in terms of L, a, b color space conventions,21 L (positive represents white

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and 0 represents dark), a (positive is red and negative is green), and b (positive is yellow and negative is

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blue). Water activity, aw, of dried PC was analyzed in duplicate using a chilled-mirror dew point sensor

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water activity meter (AquaLab Model CX-1, Decagon Devices Inc., Pullman, WA, USA). Tray dried

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PC, was submitted to the Experiment Station Chemical Laboratories, University of Missouri, USA for 8 ACS Paragon Plus Environment

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crude fat (ether extract) (AOAC official method 920.39 A), ash (AOAC official method 942.05), crude

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fiber (AOAC official method 978.10), acid detergent fiber (AOAC official method 973.18 A−D),

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neutral detergent fiber,22 amino acid profile analysis (AOAC official method 982.30 a−c), lysine

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availability (AOAC official method 975.44), phosphorus, (AOAC official method 966.01), sulfur

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(AOAC official method 956.01), calcium, sodium, magnesium, selenium, potassium, iron, zinc, copper,

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manganese, chromium, arsenic, and cadmium (AOAC official method 990.08). These analyses were

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conducted in duplicate.

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Statistical Analysis. Analysis was conducted and where applicable analysis of variance

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(ANOVA) was utilized for mean comparisons. Duncan’s multiple-range test was performed using the

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SPSS program (SPSS 21.0, IBM Corp., Armonk, NY, USA). A P value of 0.05 was used as the level of

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significance.

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RESULTS AND DISCUSSION

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Production of PC: Laboratory Scale. Slurry I is a mixture of entrained solution, similar in

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composition to liquid I, and protein rich particles. As the concentration of protein (db) is higher in slurry

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I than liquid I, it is reasonable to assume that washing slurry I, to remove the entrained solution

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surrounding the particles, might produce a product that has greater protein content (db) than dried slurry

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I (db). The sedimentation rate of particles is increased with centrifugation14 and some water surrounding

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the particles might be removed by centripetal force generated during centrifugation. This would suggest

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that slurry produced by centrifugation and not settling might have higher protein content (db) than slurry

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I prepared by gravity sedimentation. Therefore, centrifuging and washing slurry might improve protein

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content for use as an animal feed. In addition, solution surrounding particles might be recovered for

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subsequent solute separation. Slurry I protein content was 46 to 48% (w/w, db) for replicate 1 and 212

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but this was improved to between 58 and 62% (w/w, db) for replicate 1 (Figure 3) and 2 (Figure S1 of

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the Supporting Information), respectively by centrifuging and washing. These results indicated that 9 ACS Paragon Plus Environment


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separating particles from fermentation medium after TSF by centrifugation and washing produces a

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product with greatly improved protein concentration.

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The organic solutes present in liquid I, 1,3-PD, acetic acid, glycerol, GPC, and lactic acid, were also

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present in wash water at reduced concentration (Figure 4 and Figure S2 of the Supporting Information).

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These trends agreed with Steffan et al.15 who utilized a washing process to recover bacterial cells and

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sediment from soil samples. They noted that the yield of non-bacterial (organic material) was improved

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by washing the cell pellet. Wash water fractions (liquid II−liquid V) can be combined with liquid I from

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TSF and might be utilized as a source of soluble compounds. It was discovered that repeated washing

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produced very dilute streams of compounds especially after three washes. Recovering compounds from

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such dilute streams is unlikely to occur in an industrial process as the volume of solution would increase

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with additional washes. Moreover, organic solute concentrations in the 2nd and 3rd wash did not

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substantially improve compound yield (Figure 4 and Figure S2 of the Supporting Information).

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Therefore, two washing treatments were used for recovering compounds from pilot-scale processing.

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Production of Wet PC. The mass balance of slurry I from replications of small-scale TSF and solid

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and liquid fraction from washing were recorded (Figure 2 for replicate 1 and Table S1 of Supporting

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Information for replicate 2). Protein content of solids (solid III decanter) after two washing treatments

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improved to 63−64% (w/w, db) from 47−49% (w/w, db) in slurry I at the end of TSF (Table 2 and Table

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S2 of Supporting Information). Trends in protein content were similar to those observed when washing

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slurry I in laboratory scale process. Centrifugation could press liquid surrounding particles from solids

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and removed entrained solution. Solutes in liquid surrounding particles were removed producing a solid

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with elevated protein content (db) when compared with slurry prepared by gravity sedimentation.

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Moreover, it is likely that washed solids (PC), would be useful as animal feed ingredients because of

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their elevated protein content. However, conditions for drying this solid after washing should be

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explored to determine if this product has utility as an animal feed ingredient. Chemical analyses


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including amino acid profile, and lysine availability, of dried PC would provide useful preliminary

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information on the quality of this material in diets.

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As expected, both centrifugation and washing reduced the concentration of organic solutes associated

240

with particles and the concentration of organic solutes from the 2nd wash were lower than that of the 1st

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wash, and 1st pass, in that order (Figure 5 and Figure S3 of Supporting Information). Therefore, the

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majority of valuable compounds were recovered from slurry I by centrifugation and washing. The liquid

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fractions obtained from washing could be combined with liquid I from TSF as a raw material for

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compound extraction. Pilot-scale washing studies of slurry I confirmed that centrifugation and washing

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could be repeated and scaled-up using industrial equipment.

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Filtration of Liquid I, Liquid I Desludger, Liquid II Desludger, and Liquid III Desludger. Mass

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balance and microfiltration flux of a cross-flow microfiltration cell operating on decanted and washed

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liquids (liquid I, liquid I desludger, liquid II desludger, and liquid III desludger) was studied (Table 3).

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Transmembrane flux was constant, while the majority of liquid from TSF and washing passed through

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the membrane. This indicated that membrane fouling and membrane polarization were minimal or did

251

not occur and these solutions were not colloidal. Solutions had a low content of particles with sizes

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greater than 0.2 µm. The tendency of protein content and the concentration of organic solutes in the

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microfilter retentate were higher than in the permeate when filtering liquid I from TSF and liquid from

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washing slurry I (Tables 4 and 5). The higher concentration of organic compounds in retentate

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compared to those in permeate could be explained as the entrained liquid surrounding the particles

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present in retentate. The majority of protein in liquid streams was concentrated by filtration and

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recovered in the retentate. In addition, organic solutes were also partially concentrated by filtration.

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Specifically, the concentration of 1,3-PD and acetic acid in permeate and retentate from filtering liquid I

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desludger was higher than those in permeate and retentate from filtering liquid I from TSF. It is possible

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that microorganisms in slurry I are actively metabolising glycerol and lactic acid, while liquid I is not in

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contact with these bacteria as stated by Ratanapariyanuch et al.12 Therefore, the concentration of 1,3-PD 11 ACS Paragon Plus Environment


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and acetic acid in liquid I desludger was higher than that of in the liquid I from TSF due to metabolism

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by bacterial concentrates. The concentration of 1,3-PD and acetic acid precursor molecules, glycerol and

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lactic acid, in liquid I desludger was lower than those in liquid I from TSF. This further confirms the

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ability of microbial biomass associated with particles to convert glycerol and lactic acid to 1,3-PD and

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acetic acid, respectively.

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Mass balance, protein balance, and balance of individual compounds from pilot-scale processing

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were calculated (Figure 2 and Figures S4−S9 of the Supporting Information). The data indicate that

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mass, protein, and compound losses occurred in all steps in the process from fermentation until filtration

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of liquid products. Within the margin of error, there was no loss of mass in the separation of slurry I and

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liquid I. The results showed that the percent recovery of mass, protein, glycerol, 1,3-PD, lactic acid,

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acetic acid, and GPC were 96, 66, 76, 72, 77, 74, and 84%, respectively.

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Drying PC. Spray drying PC water mixtures at an inlet temperature of 149 °C resulted in outlet

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temperature of 90 °C. Heat energy was transferred to PC, which was sprayed in the drying column to

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evaporate water and the outlet temperature was, thereby, reduced. However, using an outlet temperature

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of 90 °C allowed PC solution to be dried to a powder in a single step and accumulate in the dried

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product reservoir. Chaubal and Popescu23 reported excessive collection of particles in the spray chamber

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when operating a spray drier at higher inlet temperatures. In contrast, the lower inlet temperature may

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result in unacceptably high product moisture content. A major concern when drying PC and PC water

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mixtures is the potential for heat degradation of protein and lysine. Consequently, lower inlet

281

temperatures of 139 and 129 °C were studied. Spraying at these temperatures resulted in outlet

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temperatures of 83 and 75 °C, respectively. An outlet temperature of 75 °C was sufficient to dry the PC

283

water mixture to a powder. Production cost for drying PC with lower temperature spray drying would be

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advantageous.

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Drum drying was also tested. With a drum rotation speed setting of 2, the temperature at the drum

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surface was 110 °C and the wet PC was not fully dried. Apparently, higher drum rotation speed did not 12 ACS Paragon Plus Environment

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allow sufficient time for drying. Therefore, drum rotation speed was reduced to setting number 1 to

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achieve a higher surface temperature (130 °C) and longer contact time as discussed by Valous et al.24

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The wet PC was dried acceptably under these conditions.

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PC was tray dried by moisture diffusion. The moisture is transported to the surface to be evaporated

291

as discussed by Doymaz (2004).25 The temperature for drying was maintained at 70 °C to avoid damage

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to lysine. The PC remained in the oven until samples were dry.

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Samples of spray dried, drum dried, and tray dried PC were analyzed to determine protein and

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moisture content. The products were similar with 60% (w/w, db) protein and 3−6% moisture (Table 6).

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Based on dried PC protein content and the results from 16S ribosome sequencing,12 dried PC could be

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utilized as the protein source for animal feed to replace soybean meal and fishmeal. In addition, it could

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be considered as a safe protein source for animal feed, as L. panis, L. helveticus, and L. gallinarum

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contributed most sequences that were observed in slurry I. Moreover, animals fed PC might benefit if

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the lactobacilli in the wet slurry are probiotic.13 However, dried products might not contain live bacteria.

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Furthermore, because of its low moisture content, it may be stored at ambient temperature and should

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provide long shelf-life.

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Drying method significantly affected dried PC color (P < 0.05) (Table 7). Non-enzymatic browning,

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i.e. Maillard reactions, could occur due to reactions between primary amines and reducing sugars during

304

drying. Accumulation of Maillard reaction products is a function of aw, the nature of amino acid

305

compounds and reducing sugar, pH, temperature, and time. Temperature and time promote the

306

accumulation of Maillard reaction products.26 The drum dryer temperature (130 °C) was higher than the

307

spray dryer temperature (75 °C) and tray dryer temperature (70 °C). The dried PC produced by drum

308

drying was darker, redder, and yellower compared to dried PC produced by spray and tray drying

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(Figure 6 and Table 7). The product produced by spray drying was lighter, less red and yellow than the

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tray dried product. Even though the spray drying temperature was slightly higher than the drying



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temperature of the tray dryer, the drying time was much shorter (3−4 days). Therefore, there was less

312

accumulation of Maillard reaction products in spray dried PC.

313

Water, specifically aw, is an important factor associated with food stability and degradation as

314

hydrolytic chemical reactions and microbial growth increase with aw. Therefore, safety, stability, and

315

other properties can be predicted from aw.27 All dried PC had low aw (Table 8). Nevertheless, the PC

316

drying method significantly affected aw (P < 0.05). It was discovered that PC produced by tray drying

317

had the highest aw followed by drum dried and spray dried products. The temperature utilized for tray

318

drying was low compared to temperatures for spray and drum drying. The aw of dried PC produced by

319

spray drying was slightly lower than aw of dried PC produced by drum drying. These observations could

320

be explained by spray dryer efficiency. The aw of dried PC was less than 0.5. The products are

321

sufficiently dry to prevent bacterial growth and to achieve a long storage life. In addition, Fennema et

322

al.28 stated that most reaction rates tended to decrease when aw was below 0.75−0.85. Low aw could

323

benefit dried PC storage life by increasing safety and shelf life.

324

Tray dried PC was analyzed for its nutrient composition (Table 9). The crude protein content was

325

comparable to soy protein concentrate and much higher than that of western wheat (parent grain). Crude

326

fat in dried PC was higher than either western wheat or soy protein concentrate. Similarly essential

327

amino acids present in dried PC were higher than those in western wheat. Glutamic acid was the major

328

amino acid in dried PC much like wheat29; however, dried PC contained lower lysine than soy protein

329

concentrate. Therefore, lysine should be supplemented when dried PC is used as an animal feed; lysine

330

plays a major role in building muscle and supporting body weight gain.30 Dried PC had excellent lysine

331

availability 1.82 g/100 g of protein or 95%. In addition, the lysine availability of dried PC was higher

332

than lysine availability of wheat protein (80−89%).31 The dried PC had high sulfur content potentially

333

due to the presence of sulfur-containing amino acids (cysteine and methionine). Moreover, dried PC had

334

low metal content (arsenic, cadmium, chromium, and selenium, Table 9) and, in terms of heavy metal

335

content, would be considered a safe animal feed. 14 ACS Paragon Plus Environment

Page 15 of 39


336

In conclusion, TSF slurry had high protein content (50%, w/w, db), which was more concentrated

337

than W-TS protein (38−43%, w/w, db). Due to the relatively high protein content and safe microbial

338

constituents, it is likely that slurry I would provide excellent utility as a protein source in animal feed.

339

However, liquid surrounding particles entrains organic solutes in slurry I. Therefore, washing was

340

studied to recover these solutes from TSF and improve protein content. The results from laboratory

341

processing suggested that washing twice was sufficient to recover most organic solutes from the slurry.

342

Fortuitously, the solids recovered after washing contained 60% protein (w/w, db). Moreover, tests using

343

pilot-scale equipment showed that washing is readily scaled-up to produce a 60% protein product at an

344

industrial level. Furthermore, the liquids from TSF and slurry washing fractions were readily filtered

345

through a crossflow microfiltration device with 0.2 µm membrane without indication of flux drop or

346

membrane fouling. The mass and protein balance of solids from TSF, washing, and filtration processes

347

indicated 4% of total mass loss and 34% protein loss. In addition, organic solute recovery included

348

glycerol (76%), 1,3-PD (72%), lactic acid (77%), acetic acid (74%), and GPC (84%). The PC produced

349

from pilot-scale processing was dried using a spray dryer, drum dryer, or tray dryer. Dried PC produced

350

from these 3 methods had low moisture content (4−6%) and aw (0.3). This suggested that chemical

351

reactions and microbial growth will be very slow during storage of these dried products. This property

352

would benefit PC storage and shipping. The PC from tray drying had high glutamic acid and low lysine

353

contents even though the lysine availability (95%) was excellent. Lysine should be supplemented prior

354

to use of PC as a protein source in animal feed. In addition, PC had very low heavy metal content.

355

ASSOCIATED CONTENT

356

Supporting Information

357

Supplemental tables: Mass balance of clarification of replications of small-scale TSF replicate 2 (Table

358

S1) and Protein and moisture contents of wash fractions from pilot-scale processing replicate 2 (Table

359

S2), Supplemental figures: Moisture and protein contents (A) moisture content of wash fractions 15 ACS Paragon Plus Environment


Page 16 of 39

360

fermenter 1, (B) protein content of wash fractions fermenter 1, (C) moisture content of wash fractions

361

fermenter 1, and (D) protein content of wash fractions fermenter 2 from washing slurry I from

362

fermentation at 25 °C replicate 2 (Figure S1), Concentration (g/L) of organic solutes of liquid fractions

363

from washing slurry I from fermentation at 25 °C of (A) fermenter 1 and (B) fermenter 2 replicate 2

364

(Figure S2), Concentration (g/L) of 1,3-PD, acetic acid, glycerol, GPC, and lactic acid from washing of

365

replications of small-scale TSF of (A) slurry I, (B) liquid after decanting, and (C) liquid after desludging

366

replicate 2 (Figure S3), Protein balance of pilot-scale processing replicate 1 (Figure S4), Glycerol

367

balance of pilot-scale processing replications of small-scale TSF replicate 1 (Figure S5), 1,3-PD balance

368

of pilot-scale processing replications of small-scale TSF replicate 1 (Figure S6), Lactic acid balance of

369

pilot-scale processing replicate 1 (Figure S7), Acetic acid balance of pilot-scale processing replications

370

of small-scale TSF replicate 1 (Figure S8), and GPC balance of pilot-scale processing replications of

371

small-scale TSF replicate 1 (Figure S9). This material is available free of charge via the Internet at

372

http://pubs.acs.org.

373

AUTHOR INFORMATION

374

Corresponding Authors

375

*Tel: +1 306 9665050; Fax: +1 306 9665015; E-mail: [email protected].

376

*Tel: +1 306 9665027; E-mail: [email protected].

377

Funding

378

This research was supported by the Strategic Research Program, Agricultural Development Funds of the

379

Saskatchewan Ministry of Agriculture (Grants 20080204 and 20140277), and Feeds Opportunities from

380

the Biofuels Industries Network.

381

Notes

382

The authors declare no competing financial interest.


Page 17 of 39


383

ACKNOWLEDGEMENTS

384

The authors acknowledge Pound-Maker Agventures Ltd. (Lanigan, SK, Canada) for kindly supplying

385

W-TS. POS Bio-Sciences (Saskatoon, SK, Canada) is recognized for kindly help in pilot-scale

386

processing.



387

Page 18 of 39

REFERENCES

388

(1) Meredith, J. Dryhouse design: Focusing on reliability and return on investment, In The alcohol

389

textbook, 4th ed.; Jacques, K. A., Lyons, T. P., Kelsall, D. R., Eds.; Nottingham University Press:

390

Nottingham, UK, 2003; pp 363–376.

391 392

(2) Ratanapariyanuch, K. Protein extraction from mustard (Brassica juncea L. Czern.) meal using thin stillage. M.Sc. Thesis. University of Saskatchewan, Saskatoon, SK, Canada, 2009.

393

(3) Ratanapariyanuch, K.; Shen, J.; Jia, Y.; Tyler, R. T.; Shim, Y. Y.; Reaney, M. J. T. Rapid NMR

394

method for the quantification of organic compounds in thin stillage. J. Agric. Food Chem. 2011, 59,

395

10454–10460.

396

(4) Ingledew, W. M.; Austin, G. D.; Kelsall, D. R.; Kluhspies, C. The alcohol industry: How has it

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changed and matured. In The alcohol textbook, 4th ed.; Ingledew, W. M., Kelsall, D. R., Austin, G. D.,

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Kluhspies, C., Eds.; Nottingham University Press: Nottingham, UK, 2009; pp 1–6.

399

(5) Hoppe,

K.

Sources

and

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North

Dakota.

400

https://www.ag.ndsu.edu/cattledocs/backgrounding-cattle-breakeven-calculations/files/fileinnercontentproxy.2010-

401

10-15.8952439134 (accessed on August 20, 2016).

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(6) Wilkins, M. R.; Singh, V.; Belyea, R. L.; Buriak, P.; Wallig, M. A.; Tumbleson, M. E.; Rausch,

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K. D. Effect of pH on fouling characteristics and deposit compositions in dry-grind thin stillage. Cereal

404

Chem. 2006, 83, 311–314.

405

(7) Ham, G. A.; Stock, R. A.; Klopfenstein, T. J.; Larson, E. M.; Shain, D. H.; Huffman, R. P. Wet

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corn distillers byproducts compared with dried corn distillers grains with solubles as a source of protein

407

and energy for ruminants. J. Anim. Sci. 1994, 72, 3246–3246.

408

(8) Mustafa, A. F.; McKinnon, J. J.; Ingledew, M. W.; Christensen, D. A. The nutritive value for

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ruminants of thin stillage and distillers' grains derived from wheat, rye, triticale and barley. J. Sci. Food

410

Agric. 2000, 80, 607–613.

411 18 ACS Paragon Plus Environment

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(9) Belyea, R.; Eckhoff, S.; Wallig, M.; Tumbleson, M. Variability in the nutritional quality of distillers solubles. Bioresour. Technol. 1998, 66, 207–212. (10) ADM. Feed ingredients catalog. http://www.adm.com/en-US/products/Documents/ADM-FeedIngredients-Catalog.pdf, accessed on July 18, 2016. (11) ADM.

Soycomil-R.

http://www.adm.com/en-US/Products/_layouts/ProductDetails.aspx?

productId=430 (accessed on July 20, 2016).

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(12) Ratanapariyanuch, K.; Shim, Y. Y.; Emami, S.; Reaney, M. J. T. Production of protein

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concentrate and 1,3-propanediol by wheat-based thin stillage fermentation. J. Agric. Food Chem. 2016,

420

Manuscript ID: jf-2016-05114m.

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(13) Pedersen, C.; Jonsson, H.; Lindberg, J. E.; Roos, S. Microbiological characterization of wet

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wheat distillers' grain, with focus on isolation of lactobacilli with potential as probiotics. Appl. Environ.

423

Microbiol. 2004, 70, 1522–1527.

424

(14) Geankoplis, C. J. Mechanical-physical separation processes In Transport processes and

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separation process principles (Includes unit operations), 4th ed.; Prentice Hall Professional Technical

426

Reference: Upper saddle river, USA, 2003, pp 903–954.

427 428 429 430 431 432

(15) Steffan, R. J.; Goksøyr, J.; Bej, A. K.; Atlas, R. M. Recovery of DNA from soils and sediments. Appl. Environ. Microbiol. 1988, 54, 2908–2915. (16) Ratanapariyanuch, K. Recovery of protein and organic compounds from secondary-fermented thin stillage. Ph.D. Thesis. University of Saskatchewan, Saskatoon, SK, Canada, 2016. (17) Reaney, M. J. T.; Haakensen, M.; Korber, D.; Tanaka, T.; Ratanapariyanuch, K. Process for the conversion of glycerol to 1,3-propanediol. US Patent Application 2013/0316417 A1, 2013.

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(18) Ingledew, W. M. Yeast stress in the fermentation. In The alcohol textbook, 4th ed.; Ingledew, W.

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M., Kelsall, D. R., Austin, G. D., Kluhspies, C., Eds.; Nottingham University Press: Nottingham, UK,

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2009; pp 115–126.



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(19) AOAC. Official Methods of Analysis, 18th ed.; Horwitz, W., Latimer, G. W., Eds.; Association of Official Analytical Chemists: Gaithersburg, MD, 2006. (20) Sosulski, F. W.; Imafidon, G. I. Amino acid composition and nitrogen-to-protein conversion factors for animal and plant foods. J. Agric. Food Chem. 1990, 38, 1351–1356.

440

(21) Hunter, R. S. Photoelectric color-difference meter. J. Opt. Soc. Am. 1958, 48, 985–995.

441

(22) Holst, D. O. Holst filtration apparatus for vansoest detergent fiber analyses. J. AOAC Int. 1973,

442 443 444 445 446 447 448

56, 1352–1356. (23) Chaubal, M. V.; Popescu, C. Conversion of nanosuspensions into dry powders by spray drying: a case study. Pharm. Res. 2008, 25, 2302–2308. (24) Valous, N. A.; Gavrielidou, M. A.; Karapantsios, T. D.; Kostoglou, M. Performance of a double drum dryer for producing pregelatinized maize starches. J. Food Eng. 2002, 51, 171–183. (25) Doymaz, I. Convective air drying characteristics of thin layer carrots. J. Food Eng. 2004, 61, 359–364.

449

(26) BeMiler, J. N.; Huber, K. C. Carbohydrats, In Fennema's food chemistry, 4th ed.; Damodaran,

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S., Parkin, K. L., Fennema, O. R., Eds.; Taylor & Francis Group, Boca Raton, FL, USA, 2008; pp 83–

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154.

452

(27) Reid, D. S.; Fennema, O. R. Water and ice. In Fennema's food chemistry, 4th ed.; Damodaran,

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S., Parkin, K. L., Fennema, O. R., Eds.; Taylor & Francis Group, Boca Raton, FL, USA, 2008; pp 17–

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82.

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(28) Fennema, O. R., Demodaran, S.; Parkin, K. L. Introduction to food chemistry. In Fennema's

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food chemistry, 4th ed.; Damodaran, S., Parkin, K. L.; Fennema, O. R., Eds.; Taylor & Francis Group,

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Boca Raton, FL, USA, 2008; pp 1–14.

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(29) Woychik, J. H.; Boundy, J. A.; Dimler, R. J. Wheat gluten proteins, amino acid composition of proteins in wheat gluten. J. Agric. Food Chem. 1961, 9, 307–310.


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460 461 462 463 464 465


(30) Tesseraud, S.; Larbier, M.; Chagneau, A. M.; Geraert, P. A. Effect of dietary lysine on muscle protein turnover in growing chickens. Reprod. Nutr. Dev. 1992, 32, 163–175. (31) Sarwar, G.; Bowland, J. P. Availability of amino acids in wheat cultivars used in diets for weanling rats. Can. J. Anim. Sci. 1975, 55, 579–586. (32) Bell, B. Wheat for animal feed. http://www.omafra.gov.on.ca/english/livestock/beef/facts/ wheat.htm (accessed on July 21, 2016).

466



Page 22 of 39

467

FIGURE CAPTIONS

468

Figure 1. Flowchart of small-scale washing process.

469

Figure 2. Mass balance of pilot-scale processing.

470

Figure 3. Moisture and protein contents (A) moisture content of fermenter 1 wash fractions, (B) protein

471

content of fermenter 1 wash fractions, (C) moisture content of fermenter 1 wash fractions, and (D)

472

protein content of fermenter 2 wash fractions after washing slurry I from fermentation at 25 °C

473

(replicate 1). Each value is presented as the mean ± standard deviation (SD, n = 2). Total nitrogen was

474

determined by the Kjeldahl method. Corrected protein was calculated using conversion factor of 5.7

475

times N and expressed as crude protein. Nitrogen in the samples contributed by GPC and betaine was

476

determined by DPFGSE-NMR. Nitrogen contributed by these materials to total nitrogen was subtracted

477

prior to calculation of protein content.

478

Figure 4. Concentration (g/L) of organic solutes of liquid fractions from washing slurry I after

479

fermentation at 25 °C of (A) fermenter 1 and (B) fermenter 2 replicate 1. Due to interfering resonances

480

from carbohydrate and protein the concentration of glycerol might be overestimated.

481

Figure 5. Concentration (g/L) of 1,3-PD, acetic acid, glycerol, GPC, and lactic acid from washing of

482

replications of small-scale TSF of (A) slurry I, (B) liquid after decanting, and (C) liquid after desludging

483

replicate 1.

484

Figure 6. Images of protein concentrate produced by; (A) spray drier, (B) drum drier, and (C) tray drier.


Page 23 of 39


485

Table 1. Feed Ratea of Decanter and Desludger of the 1st Pass, 1st Wash, and 2nd Wash for

486

Washing Slurry I from Replications of Small-scale TSF Replicate 1 and 2 feed rate (kg/h)

replicate 1

replicate 2

decanter

desludger

decanter

desludger

st

300

320

300

300

st

400

400

350

350

1 pass 1 wash nd

2 wash 400 400 400 400 Feed rate of decanter and desludger was determined based on the ability of the centrifuge to prevent

487

a

488

particle bypass into the supernatant.

489



490

Page 24 of 39

Table 2. Protein and Moisture Contentsa of Wash Fractions from Pilot-scale Processing sample

proteinb (%, w/w, wb)

moisture (%, w/w, wb)

protein (%, w/w, db)

slurry Ic

4.54 ± 0.00

90.30 ± 0.00

46.79 ± 0.05

10.91 ± 0.06

82.03 ± 0.04

60.69 ± 0.18

0.88 ± 0.00

96.61 ± 0.00

25.94 ± 0.02

2.52 ± 0.05

93.07 ± 0.02

36.29 ± 0.70

0.72 ± 0.01

96.95 ± 0.01

23.63 ± 0.30

3.89 ± 0.07

92.96 ± 0.00

55.17 ± 1.03

solid I decanter liquid I decanter

c

solid I desludger liquid I desludger

c st

c

mixture of solid and water before 1 wash solid II decanter

11.29 ± 0.07

82.37 ± 0.02

64.04 ± 0.32

c

0.70 ± 0.00

98.05 ± 0.01

35.74 ± 0.28

solid II desludger

1.94 ± 0.01

95.74 ± 0.03

45.51 ± 0.50

liquid II desludgerc

0.32 ± 0.02

98.63 ± 0.04

23.47 ± 1.02

mixture of solid and water before 2nd washc

3.39 ± 0.05

94.18 ± 0.00

58.30 ± 0.94

10.27 ± 0.00

83.60 ± 0.06

62.64 ± 0.23

0.32 ± 0.01

99.15 ± 0.01

37.90 ± 0.69

0.59 ± 0.01

98.60 ± 0.02

42.47 ± 0.19

liquid II decanter

solid III decanter liquid III decanter

c

solid III desludger c

liquid III desludger 0.17 ± 0.01 99.30 ± 0.01 24.20 ± 0.41 b Each value is presented as the mean ± SD (n = 2). Total nitrogen was determined by the Kjeldahl

491

a

492

method. Corrected protein was calculated using conversion factor 5.7 and expressed as crude protein.

493

c

494

by these materials to total nitrogen was subtracted prior to calculation of the protein content.

Nitrogen contributed by GPC and betaine was determined by DPFGSE-NMR. The nitrogen contributed


Page 25 of 39

495


Table 3. Mass Balance from Filtration Replications of Small-scale TSF weight of liquid (kg)

weight of permeate (kg)

weight of retentate (kg)

hour of operating filter unit

flux (L/m2/h)

liquid I from TSF

66.28

63.09

3.31

1.41

75

liquid I desludger

140.88

129.82

10.17

3.83

56

liquid II desludger

103.56

94.06

9.31

3.42

46

liquid III desludger

116.78

99.55

7.30

3.67

45

sample

496



Page 26 of 39

497

Table 4. Protein and Moisture Contentsa of Filtration Fractions from Replicates of Small-scale

498

TSF proteinb (%, w/w, wb)



liquid I from TSF

0.72 ± 0.03

96.76 ± 0.00

22.15 ± 0.12

permeate of liquid I after filtering

0.41 ± 0.02

97.52 ± 0.01

16.49 ± 0.54

retentate of liquid I after filtering

2.31 ± 0.03

92.37 ± 0.03

30.31 ± 0.30

liquid I desludger

0.72 ± 0.00

96.95 ± 0.03

23.63 ± 0.30

permeate of liquid I desludger after filtering

0.34 ± 0.01

97.70 ± 0.00

14.60 ± 0.47

retentate of liquid I desludger after filtering

2.51 ± 0.02

90.20 ± 0.02

25.64 ± 0.20

liquid II desludger

0.32 ± 0.02

98.63 ± 0.04

23.47 ± 1.02

permeate of liquid II desludger after filtering

0.08 ± 0.01

99.90 ± 0.00

8.25 ± 1.17

retentate of liquid II desludger after filtering

1.02 ± 0.00

96.52 ± 0.02

29.35 ± 0.06


0.17 ± 0.01

99.30 ± 0.01

24.20 ± 0.41

permeate of liquid III desludger after filtering

0.01 ± 0.00

99.49 ± 0.01

2.69 ± 0.50

sample

retentate of liquid III desludger after filtering 0.76 ± 0.01 97.82 ± 0.20 34.79 ± 0.62 b Each value is presented as the mean ± SD (n = 2). Total nitrogen was determined by the Kjeldahl

499

a

500

method. Corrected protein was calculated using conversion factor 5.7 and expressed as crude protein.

501

c

502

by these materials to total nitrogen was subtracted prior to calculation of the protein content.

Nitrogen contributed by GPC and betaine was determined by DPFGSE-NMR. The nitrogen contributed


Page 27 of 39


503

Table 5. Concentration (g/L) of 1,3-PD, Acetic Acid, Glycerol, GPC, and Lactic acid of Filtration Fractions from Replications of Small-

504

scale TSF compound

liquid I

liquid I desludger

liquid II desludger


permeate

retentate

permeate

retentate

permeate

retentate

permeate

retentate

4.72b

5.44

5.68

5.95

2.73

2.98

1.47

1.56

4.84

5.59

5.51

5.68

2.62

2.98

1.59

1.58

glycerol

4.29

6.65

2.07

4.11

1.06

1.59

0.47

0.85

GPC

1.17

1.50

1.14

1.33

0.55

0.70

0.34

0.34

1,3-PD acetic acid a

lactic acid 3.87 4.52 2.27 3.21 1.33 1.70 0.73 0.92 b The concentration of glycerol in the samples may be affected by the presence of interfering resonances from carbohydrate and protein. Data are

505

a

506

the means of duplicate analysis.

507



508

Page 28 of 39

Table 6. Protein and Moisture Contentsa of Dried PC using Different Devices sample

proteinb (%, w/w, wb)



spray dryer

58.54 ± 1.32

3.86 ± 0.04

60.89 ± 1.37

drum dryer

58.11 ± 1.21

3.52 ± 0.11

60.23 ± 1.29

tray dryer 58.87 ± 0.96 5.76 ± 0.05 62.47 ± 0.99 b Each value is presented as the mean ± SD (n = 2). Total nitrogen was determined by the Kjeldahl

509

a

510

method. Corrected protein was calculated using conversion factor 5.7 as expressed as crude protein.

511


Page 29 of 39

512


Table 7. Colora of Dried PC using Different Devices sample

L

a a

spray dryer

66.61 ± 0.19

drum dryer

54.75 ± 0.17c

b c

18.22 ± 0.20c

10.36 ± 0.09a

24.82 ± 0.80a

6.78 ± 0.09

513

tray dryer 56.40 ± 0.07b 10.16 ± 0.03b 23.67 ± 0.30b a Each value is presented as the mean ± SD (n = 2). Values followed by different letters are significantly

514

different (P < 0.05).

515



516

Page 30 of 39

Table 8. awa of Dried PC at 22 °C using Different Devices drying method

aw

spray dryer

0.23 ± 0.00c

drum dryer

0.25 ± 0.00b

517

tray dryer 0.30 ± 0.01a a Each value is presented as the mean ± SD (n = 2). Values followed by different letters are significantly

518

different (P < 0.05).

519


Page 31 of 39


520

Table 9. Nutrients of Dried PCa from Tray Drying Method Comparing to Western Wheat and Soy

521

Protein Concentrate nutrient (%, w/w)

western wheatb

dried PC

soy protein concentratec

522

dry matter 94.21 ± 0.13 NAd 91.0 crude protein 59.97 ± 0.02 15.1 63.0 crude fat 6.37 ± 0.21 1.50 0.6 ash 1.57 ± 0.11 NA 6.5 crude fiber 7.22 ± 0.44 2.50 4.0 acid detergent fiber 32.94 ± 1.10 NA NA neutral detergent fiber 20.80 ± 1.19 NA NA amino acid (% as is) taurine 0.04 ± 0.01 NA NA hydroxyproline 0.01 ± 0.01 NA NA aspartic acid 3.25 ± 0.01 NA NA threonine 2.08 ± 0.01 0.49 2.73 serine 2.70 ± 0.04 NA NA glutamic acid 17.73 ± 0.21 NA NA proline 6.04 ± 0.08 NA NA lanthionine 0.00 ± 0.00 NA NA glycine 2.28 ± 0.01 NA NA alanine 2.39 ± 0.01 NA NA cysteine 1.44 ± 0.00 0.34 0.98 valine 3.19 ± 0.04 0.84 3.38 methionine 1.22 ± 0.00 0.24 0.91 isoleucine 2.60 ± 0.00 0.72 3.19 leucine 4.94 ± 0.00 1.34 5.20 tyrosine 1.97 ± 0.00 NA NA phenylalanine 3.17 ± 0.01 0.91 3.45 hydroxylysine 0.05 ± 0.00 NA NA ornithine 0.07 ± 0.01 NA NA lysine 1.92 ± 0.01 0.45 4.23 histidine 1.51 ± 0.01 NA 1.82 arginine 2.92 ± 0.00 0.69 4.94 tryptophan 0.64 ± 0.01 0.15 0.78 available lysine 1.82 ± 0.01 NA NA a b 32 c Each value is presented as the mean ± SD (n = 2). Wheat for animal feed, Feed ingredients10 and

523

Soycomil-R11 d NA = not available.

524



Page 32 of 39

525

Table 9. Nutrients of Dried PCa from Tray Drying Method Comparing to Western Wheat and Soy

526

Protein Concentrate (Cont’d) dried PC

western wheatb

soy protein concentratec

phosphorus

0.35 ± 0.00

NAd

0.8

sulfur

0.80 ± 0.14

0.18

NA

calcium

0.20 ± 0.00

0.04

0.35

sodium

0.04 ± 0.00

0.03

0.01

magnesium

0.04 ± 0.00

0.36

0.32

potassium

0.13 ± 0.00

0.40

2.20

Protein Concentrate Production from Thin Stillage - Journal of

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