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Processing methods for grains with
special reference to particle size
Final report prepared for the
Co-operative Research Centre for High Integrity Australian Pork
(formerly the Co-operative Research Centre for an Internationally
Competitive Pork Industry)
By
Peter Sopade1, Mike Gidley
1 and John Black
2
1Centre for Nutrition and Food Sciences
Queensland Alliance for Agriculture and Food Innovation
The University of Queensland
St Lucia 4072
and
2John L Black Consulting
Locked Bag 21
Warrimoo 2774,
December 2011
2
Table of Contents
Executive Summary
5
A. Introduction
6
B. Research Approach
6
B.1 Sieve analysis of ileal digesta and diet
6
B.2 Survey of feed mills
7
B.3 Development of an in-vitro protein digestion procedure and modelling
protein digestograms
7
B.4 Laboratory analysis of selected commercially milled grains
9
C. Results and Discussion
10
C.1 Particle size distribution of ileal digesta and corresponding diet
10
C.2 Particle size distribution of commercially milled grains
14
C.3 Laboratory analysis of selected commercially milled grains
19
C.3.1 Evaluation of the in-vitro protein digestion procedure
19
C.3.2 Laboratory analysis of commercially milled grains
24
D. Conclusions
28
E. Further research needs and recommendations
30
Acknowledgements 30
3
List of Figures
1
Rehydration behaviours of the selected diets (A – immediately after soaking,
B – after about 16 hr of soaking)
11
2
Particle size distribution of the digesta and corresponding diets 12
3
Typical particles retained from the digesta and diets (sieve, 1000 µm) 13
4
Typical disc, roller and hammer mills with the main milling forces 14
5
Venn diagram of the distribution of the milling techniques amongst the
surveyed feed mills (values are the number of mills using the technique or a
combination of techniques)
15
6
Typical particle size distributions of the milled grains 16
7
Design of a manual sieving device 20
8
Protein digestograms of feed ingredients 22
9
Protein digestograms of processed and non-processed diverse materials 23
10
Protein and starch digestograms of the commercially-milled sorghum and
wheat
25
11
Plots of rates of starch and protein digestion, and square of geometric mean
particle size
27
12
Starch digestograms showing differences between mills 29
4
List of Tables
1
Information on and ileal digestibility of the diets studied for particle size
distribution
10
2
Particle size parameters of the milled grains from the surveyed feed mills 17
3
Approximate regular bodies for grains and the formulas for their volumes 18
4
Effect of protease treatment on in-vitro protein digestibility of feed ingredients 21
5
Effect of processing and sample diversity on in-vitro protein digestibility 21
6
Digestion and chemical properties of the commercially-milled sorghum and wheat 26
5
Executive Summary
The project demonstrated that pig ileal digesta contained large feed particles that were related
to large particulates in the diets. This emphasises the need to control feed particles to sizes that
enhance digestion and energy delivery to pigs. Towards this, the milling techniques, including
mill settings need to be chosen to keep the average particle size below a critical value (1 mm)
that is partly determined by the economics of feed processing, and for pelleted diets, partly
determined by pellet durability.
The following are the specific findings from the project:
Hammer-, roller- and disc-mills were the three main mills in the Australian feed industry.
However, even though most mills used hammer mills, more feed mills appeared to be
changing to disc mills, while some feed mills mainly mixed hammer- and roller-milled
grains to formulate their mash diets.
Particle size distributions of milled cereal grains from the Australian feed mills ranged
from narrow to broad.
Particle size distributions from wet sieving of pig diets and ileal digesta revealed that
diets with large particles could yield digesta with large particles.
An in-vitro protein digestion procedure was developed to investigate protein digestion,
and possibly the kinetics of protein digestion. This will complement the in-vitro starch
procedure developed in an earlier project, giving the Australian feed mills simple
techniques to screen feeds for digestion using in-vitro techniques..
A simple sieving device was designed for quality control during grain milling to guide
mill settings for maximum digestion and energy delivery to pigs.
Information is now available from the study on the differences in particle size distributions
between the common mills used in pig feed manufacture in Australia to highlight variability,
which may transform into pig diets and animal performance. Two methods of in-vitro
digestion, each targeting starch and protein, the main energy yielding components, are now
available for the pig industry, and they have been tested with commercial samples. While they
are not substitutes for animal experiments, the in-vitro methods are fast and can be used to
screen samples as part of a quality assurance strategy in the Australian pig industry.
With such diverse particle size distributions, future studies are to investigate the consequences
of a wide range of particle size distributions on animal performance to buttress the findings of
the in-vitro approach. The dependence of protein digestion on particle size will be further
studied. The economics of the whole feed processing is important in establishing an
appropriate particle size distribution, and how to achieve such. These issues will be addressed
in a follow-up project (4B-112 - Optimising particle size distribution for grains and protein
sources).
6
A. Introduction Previous studies indicated that digestion rate of pig feed is largely affected by milling and heat
processing. Milling is a unit operation in which particle size is reduced, and because of
differences in fracturability of various parts of a material, for example grains, a particle size
distribution is normally obtained after milling. The distribution can also occur because of the
effective milling force of a mill, and/or the nature (e.g. roughness, clearance and restriction) of
the reaction zone in the mill. In reducing the particle size of materials, the internal components
are exposed and the surface area for enzyme-substrate interactions, amongst other effects, is
increased. There are many types of mills, which include hammer-, roller-, disc-, ball-, pin, and
cryo- or freezer-mills. The effective milling forces (e.g. impact, compressive and shearing or
attrition) in these mills are different, and while some mills (e.g. hammer mill) are widely used
for diverse materials and purposes, some mills (e.g. freezer-mill) are used for specialised
products where frictional heat generation is undesirable. Also, because of process economics
(e.g. cost, throughput, mode of operation and automation), some mills are preferred for
commercial operations.
Upon grain receival and cleaning, milling is the first main manufacturing step in diet
manufacture in Australian feed mills. The situation in feed mills outside Australia is not
expected to be different. However, the predominant mills for feed manufacture in Australia
have not been documented in order to understand differences, if any, in Australian pig feeds.
Understanding this will assist explaining differences in digestibility of pig feeds in order to
map out milling strategies that maximise feed efficiency. In approaching this, the following
questions were asked in the present study:
What are the various mills used in the Australian feed industry, particularly amongst
small-scale producers where mash diets, instead of pelleted diets, dominate?
Are there differences in particle size distribution of milled grains?
How do diet particle size distributions reflect in ileal digesta particle size distributions?
What are the relationships between the rates of starch and protein digestion of
commercially-milled grains, and their particle sizes?
B. Research Approach
B.1 Sieve analysis of ileal digesta and diet
In previous studies, diet particles that escaped digestion were observed, and the particles
ranged in size. In the present study, it was thought that particle size distribution of ileal digesta
could be matched with those of corresponding diets to investigate relationships between them.
Being wet materials, wet sieving of digesta was done using sieves (Endecotts Ltd, London,
England) 1400, 1000, 710, 500, 250, and 125 µm. Shower sprays were directed at about 20 g of
manually-mixed digesta for about 5 min., and particles through the 125 µm-sieve were washed
off. The sieves were allowed to drain for about 5 min. before they were weighed. About 20 g of
7
the corresponding diets were soaked overnight (about 16 hr.) in excess water, and wet sieved as
done for digesta.
B.2 Survey of feed mills Commercial feed mills that mainly use mash diets were visited to observe their grain milling
techniques, or were contacted to obtain information about their mills. The mills in operation
were noted, and where possible, mill settings were recorded and/or requested to be changed
within the normal processing schedules. Milled grains, and whole grains, where available, were
collected.
About 100 g of milled grains were dry sieved for about 30 min. using a sieve shaker (Endecotts
shaker, ExTech Pty. Ltd., Victoria, Australia) with sieves 4000, 2800, 1400, 1000, 710, 500,
250, 125, and 75 µm. Particles through the 75 µm sieve were collected in a pan as described in
a standard procedure (American Society of Agricultural and Biological Engineers. ANSI/ASAE
S319.4 (2008). Method of determining and expressing fineness of feed materials by sieving),
which was also used to calculate geometric mean diameter (Dgw) and geometric standard
deviation of particle diameter by mass (Sgw).
The length, width and thickness of whole grains were measured using Vernier callipers, and
1000-grain weight was determined to partly describe the grains. In all, 10 commercial feed
mills were surveyed, and they are henceforth identified as Mills 1 – 10.
B.3 Development of an in-vitro protein digestion
procedure and modelling protein digestograms There are various in-vitro protein digestion procedures, which can generally be classified as
based on a single protease, such as pepsin or trypsin, or based on multi-proteases, such as
pepsin-trypsin, pepsin-pancreatin, trypsin-chymotrypsin, trypsin-chymotrypsin-peptidase,
trypsin-chymotrypsin-protease, or trypsin-chymotrypsin-peptidase-pronase (Tinus T, Damour
M, van Riel V, Sopade PA. (2012). Particle size-digestibility relationships, if starch, does
protein? Journal of Food Engineering (submitted)). These techniques either measure the
products (digested proteins) or residues (undigested proteins) of proteolysis using a batch or
continuous approach that is based on procedures, which include direct protein analysis (e.g.
Kjeldahl, Dumas or colourimetry method), amino acid analysis, chromatography, acidity (e.g.
pH drop or pH stat), and electrophoresis (e.g. sodium dodecyl sulphate polyacrylamide gel
electrophoresis, SDS-PAGE).
The pH drop three-enzyme (trypsin-chymotrypsin-peptidase or trypsin-chymotrypsin-protease)
method (Hsu HW, Vavak DL, Satterlee LD, Miller GA (1977). A multienzyme technique for
estimating protein digestibility. Journal of Food Science, 42, 1269) is widely used in
estimating apparent in-vitro protein digestibility. It involves rehydrating a sample weight that is
equivalent to 62.5 mg protein in 10 mL. of water at 37oC for 1 hr, after which the pH is
adjusted to about 8.0 with 0.1M NaOH and/or HCl before incubating with 1 mL. of a multi-
enzyme solution, which consists of about 16 mg of trypsin (Sigma T0303 Trypsin from porcine
pancreas Type IX-S, lyophilized powder, 13,000-20,000 BAEE units/mg protein), 31 mg of
chymotrypsin (Sigma C4129 α-Chymotrypsin from bovine pancreas, Type II, lyophilized
powder, ≥40 units/mg protein) and 13 mg protease (Sigma P5147 Protease from Streptomyces
8
griseus Type XIV, ≥3.5 units/mg solids). Protease from Streptomyces griseus is a substitute for
peptidase (Sigma P7500 Peptidase from porcine intestinal mucosa, 50-100 units/g solids),
which although is in the original procedure, has been discontinued. The multi-enzyme solution
is usually prepared fresh on the day of analysis and could be kept at 37oC, rather than in an ice
bath as recommended in the original procedure. The pH of the multi-enzyme solution needs to
be adjusted to about 8.0 at 37oC prior to being added to the rehydrated sample dispersion.
During method developments, reproducible results were obtained when both the rehydrated
sample dispersion and the multi-enzyme solution were maintained at 37oC and pH of about 8.0
before mixing for the digestion process. The pH of the digesta was automatically recorded
every 5 sec. for 15 min. using a programmable pH meter (smartCHEM-pH with WinTPS, TPS,
Springwood, QLD 4127). Like other in-vitro protein techniques, the original pH drop method
recommends a single-point measurement when the pH at 10 min. (pH10 min.) is used to calculate
the in-vitro protein digestibility (IVPD, %) using Eq. (1). The in-vitro protein digestibility
(100%) of casein (Sigma C7078 Casein from bovine milk) was used as the reference.
IVPD = 210.46 – 18.10 pH10 min. (1)
However, with the starting pH of 8.0, Eq. (1) can be re-written as:
IVPD = 210.46 – 18.10 (8 – ΔpH10 min.) (2)
where ΔpH10 min. is the change in pH in 10 min. from the initial pH of about 8.0.
Simplifying Eq.(2),
IVPD = 65.66 + 18.10 ΔpH10 min. (3)
Eq. (3) is more applicable because it uses the change in pH from the starting value, which
might be close to but not exactly 8.0. The limitation of Eq. (1) or Eq. (3) is that IVPD will have
a value of 65.66% with no protein digested, and consequently, no change in pH of the digesta.
Also, in the absence of enzyme inhibition or inactivation, mathematically, IVPD can have a
value that is greater than 100% when pH10min. is ≤ 6.1, or ΔpH10 min. ≥ 1.9.
From protein chemistry, the drop in pH results from the release of amino acids, protein
building units, as protein is digested. The release of amino acids during proteolysis is not
expected to be linear or of a zero order. Although a power-law model can be used to describe
protein digestograms, a modified first order kinetic can be explored to investigate and
understand the kinetics of protein digestion. In terms of pH (Eq. (4)), a modified first order
kinetic model can be written as:
pHt = pHo + pH∞-o (1 - exp [-Kpr t]) (4)
pH∞ = pHo + pH∞-o
where pHt = pH at time t, pHo = initial pH and Kpr = rate of protein digestion (min-1
). The
reciprocal of the rate of starch digestion in sorghum, for example, is directly related to the
square of the particle size of the milled grains (Mahasukhonthachat K, Sopade PA, Gidley MJ
(2010). Kinetics of starch digestion in sorghum as affected by particle size. Journal of Food
Engineering 96: 18). If Eq. (4) is found to suitably describe protein digestograms, the rate of
9
protein digestion (Kpr), can be related to particle size. As done for starch, the relationship can
be used to understand the mechanisms of protein digestion.
B.4 Laboratory analysis of selected commercially milled
grains Using milled sorghum and wheat grains from mill 7, laboratory experiments were statistically
designed for the analyses listed below. Except otherwise stated, and pending detailed statistical
analysis, means and standard deviations are presented and used in discussing the measured
trends.
a. Moisture content
Moisture content was determined using an oven method (135oC, 3 hr).
b. Protein content
The Dumas method (Waramboi JG, Dennien S, Gidley MJ, Sopade PA. 2011.
Characterisation of sweetpotato from Papua New Guinea and Australia:
Physicochemical, pasting and gelatinisation properties. Food Chemistry 126: 1759)
was used to determine the protein content (N x 6.25) of the milled grains, and
expressed on a solids basis.
c. Starch content
Starch content was determined using a procedure based on the Megazyme dimethyl
sulphoxide-α-amylase-amyloglucosidase (DMSO-AA-AMG) method, and expressed
on a solids basis.
d. In-vitro starch digestion
A glucometry method (Mahasukhonthachat K, Sopade PA, Gidley MJ (2010). Kinetics
of starch digestion in sorghum as affected by particle size. Journal of Food
Engineering 96: 18) was used, and starch digestograms were described using a
modified first order kinetic model (Eq. (5)), as discussed in previous studies, to obtain
salivary-gastric digested starch (Do, very rapidly digested starch), maximum digested
starch (D∞), rate of starch digested (Kst), area under the starch digestogram (AUC),
and digested starch at 4 hr. (D240). AUC, which is a measure of the amount of glucose
released into the blood (glycemic index), was calculated with Eq. (6).
Dt = Do + D∞-o [1 - exp (- Kst t)] (5)
where, Dt = digested starch (g/100g dry starch) at time t (min.), Do = digested starch
(g/100g dry starch) at time t = 0, D∞ = digested starch (g/100g dry starch) at infinite
time (t → ∞), and Kst = rate of starch digestion (g/min).
(6)
e. In-vitro protein digestion
The method described in B.3 was used, and protein digestograms were modelled using
Eq. (4), from which the rate of protein digestion (Kpr) was obtained.
10
C. Results and Discussion
C.1 Particle size distribution of ileal digesta and
corresponding diet The four diets that were studied, and their corresponding ileal digestibility parameters are as in
Table 1 showing digestible energy from about 9 – 12 MJ/kg. The diets varied in their
rehydration behaviours (Fig. 1), and this could be related to how they disperse in digestive
fluids. However, differences in the pH, temperature and contact time in the digestive tracts
could modify the rehydration behaviours. The diets were fully rehydrated prior to analysis
because, for comparison purposes, it was thought that the particles in the ileal digesta would
have fully rehydrated since collection during the metabolism studies that produced them.
Figure 2 shows the particle size distributions of the ileal digesta and their corresponding diets.
Generally, the diets appeared to contain larger particles than the digesta, but with no particular
trend with digestible energy (DE diet af or DE grain af). Visual observations revealed particles
in the digesta (Fig. 3), which were retained on the sieves, were mainly grain particles. The
presence of undigested starch or grain particles in ileal digesta has been highlighted in previous
studies under the current program of the Pork CRC.
The particle size distributions of the digesta studied have not revealed information to be used in
assessing their digestible energy. This could be because each digesta was collected over days
during the metabolism studies. Assuming enough quantities can be collected, the particle size
distributions of selected digesta collected at known times after feeding might provide a better
understanding of the changes in size distributions due to digestion. Follow-up studies will
investigate this on selected samples from Project 4B-112 - Optimising particle size distribution
for grains and protein sources.
Table 1: Information on and ileal digestibility of the diets studied for particle size
distribution
Pork CRC
Experiment
Code
Pork
CRC
Grain
ID No.
Pork
CRC
Diet No.
Grain Pork CRC
Information
Code
Diet Digestible
Energy As-fed
(DEaf, MJ/kg)
Grain Digestible
Energy As-fed
(DEaf, MJ/kg)
DS004 1753 22 Wheat RAW0 12.25 12.55
HR001 3897 79 Barley RAW0 8.81 8.92
HR001 6835 94 Triticale SPROUT 11.71 11.99
DS004 7714 17 Sorghum STRESS1 12.11 12.41
11
Figure 1: Rehydration behaviours of the selected diets (A – immediately after soaking, B – after about 16 hr of soaking)
Triticale
HR001D94
Sorghum
DS004D17
Barley
HR001D79
Wheat
DS004D22
Triticale
HR001D94
Sorghum
DS004D17
Barley
HR001D79
Wheat
DS004D22
A
B
12
Figure 2: Particle size distribution of the digesta and corresponding diets
0
10
20
30
40
1400 1000 710 500 250 125
Wheat
Diet DEaf = 12.25
1400 1000 710 500 250 125
Diet
Ileal
Barley
Diet DEaf = 8.81
0
10
20
30
40
1400 1000 710 500 250 125
Triticale (SPROUT)
Diet DEaf = 11.71
1400 1000 710 500 250 125
Sorghum (STRESS)
Diet DEaf = 12.71% R
etai
ned
Sieve size (µm)
13
Figure 3: Typical particles retained from the digesta and diets (sieve, 1000 µm)
Wheat - diet Wheat - digesta
Barley - diet Barley - digesta
Triticale - diet Triticale - digesta
Sorghum - diet Sorghum - digesta
14
C.2 Particle size distribution of commercially milled
grains Hammer, roller and disc mills were the main mills in the 10 feed mills surveyed. Figure 4
shows photographs of typical disc-, roller- and hammer-mills, and their main milling forces.
These forces are expected to generate differences in the particle size distributions of milled
grains, and possibly because of process requirements for specific size distributions, some feed
mills mixed milled grains from two mills on site, for example, hammer- and roller-mills (Fig.
5).
Figure 4: Typical disc-, roller- and hammer-mills with the main milling forces
15
Figure 5: Venn diagram of the distribution of the milling techniques amongst the
surveyed feed mills (values are the number of mills using the technique or a
combination of techniques)
The typical particle size distributions of grains milled in the three mills is as in Figure 6.
Although there are genotype x environmental (G x E) effects on grain fracturability, the
hammer mills appeared to generate broad distributions, the (fluted) roller mills seemed to
generate narrow distributions and the distributions with the disc mills ranged from a broad to a
skewed (fine particles) one. Because of the dominant compressive force in roller mills, some
particles in the roller-milled grains were compressed or flattened rather than broken into
particles as obtained from the other mills. Apart from resistance to enzyme transport from grain
bran or husks, compressed or flattened grains can be equally digested as broken ones because
of a reduced particle thickness. Unfortunately, the power consumed during milling in the feed
mills was not available, thereby limiting comments on the relative suitability of the mills. It is
envisaged that data on the economics of grain milling will be gathered in follow-up studies.
Table 2 shows the average particle sizes (geometric mean diameter, Dgw) and geometric
standard deviations of particle diameter by mass (Sgw) of the milled grains studied. Dgw
ranged from 0.522 – 1.093 mm for the hammer mills, 0.659 – 1.033 mm for the disc mills and
0.996 – 2.560 mm for the roller mills, while Sgw ranged from 0.450 – 0.798 mm for the
hammer mills, 0.536 – 0.882 mm for the disc mills and 0.665 – 1.235 mm for the roller mills.
16
Figure 6: Typical particle size distributions of the milled grains
Sorghum
Barley
Wheat
% R
eta
ined
Sieve size (µm)
17
Table 2: Particle size parameters of the milled grains from the surveyed feed mills*
*ns = no setting was made available
ng = no grain was collected
Diameter = hypothetical diameter of a sphere of the same volume of the grain assuming a regular shape
SRR = size reduction ratio Diameter
Dgw
Other deductions from Table 2, some of which are expected, are:
There were differences in the (ease of) fracturability of the grains with the general trend
being (maize >) sorghum > wheat > barley irrespective of the mill. This could be partly
due to the degree of toughness of the grain bran or husks.
Feed mill Grain characteristic Fracturability index
Number Type Mill setting Type Major diameter Minor diameter Thickness 1000-grain count Diameter Dgw Sgw SRR
(mm) (mm) (mm) (g) (mm) (mm) (mm)
1 Hammer 2 mm Sorghum 4.5 4.1 2.7 32 3.8 0.639 0.518 5.9
1 Roller 2 mm Sorghum 4.5 4.1 2.7 32 3.8 1.110 0.693 3.4
1 Hammer 2 mm Wheat 5.9 3.3 2.9 37 3.8 0.672 0.624 5.7
1 Roller 2 mm Wheat 5.9 3.3 2.9 37 3.8 1.170 0.873 3.3
1 Hammer 2 mm Barley 8.7 3.5 2.6 38 4.3 0.902 0.657 4.7
1 Roller 2 mm Barley 8.7 3.5 2.6 38 4.3 1.225 0.752 3.5
1 Hammer 2 mm Barley ng ng ng ng ng 0.700 0.552 ng
1 Roller 2 mm Barley ng ng ng ng ng 1.281 0.805 ng
1 Hammer 2 mm Wheat ng ng ng ng ng 0.655 0.590 ng
1 Roller 2 mm Wheat ng ng ng ng ng 1.120 0.898 ng
1 Hammer 2 mm Sorghum ng ng ng ng ng 0.590 0.452 ng
1 Roller 2 mm Sorghum ng ng ng ng ng 1.023 0.665 ng
2 Hammer ns Sorghum 4.3 3.8 2.6 30 3.6 0.722 0.533 5.0
2 Hammer ns Wheat 6.5 3.2 2.9 41 3.9 0.782 0.750 5.0
2 Hammer ns Barley 8.3 3.6 2.6 41 4.3 0.936 0.717 4.6
3 Hammer 3.2 mm Sorghum 4.6 3.8 2.5 26 3.6 1.093 0.798 3.3
3 Hammer 2.0 mm Sorghum 4.6 3.8 2.5 26 3.6 0.841 0.450 4.3
3 Ripple ns Sorghum 4.6 3.8 2.5 26 3.6 1.173 0.864 3.1
4 Roller Double Sorghum 4.8 4.2 2.9 37 4.0 1.365 0.913 2.9
4 Disc ns Sorghum 4.8 4.2 2.9 37 4.0 0.780 0.560 5.1
4 Roller Single Wheat 5.8 2.9 2.4 30 3.4 1.780 0.677 1.9
4 Disc ns Wheat 5.8 2.9 2.4 30 3.4 0.929 0.882 3.7
4 Roller Double Barley 9.0 3.4 2.5 37 4.2 1.697 0.843 2.5
4 Roller Single Maize 10.7 8.2 4.2 318 5.9 0.996 0.868 5.9
5 Hammer 3 mm Wheat 7.3 3.5 3.1 56 4.3 0.691 0.556 6.3
5 Hammer 3 mm Barley 8.7 3.5 2.5 45 4.2 0.669 0.497 6.4
6 Roller ns Maize 12.0 8.3 4.3 305 6.2 1.135 1.085 5.5
6 Roller ns Sorghum 4.8 4.0 2.8 35 3.9 1.397 1.086 2.8
6 Roller ns Barley 8.8 3.3 2.2 40 4.0 2.560 0.951 1.6
6 Roller ns Wheat 5.8 2.5 2.2 19 3.2 1.734 1.014 1.8
7 Disc 1.1 mm Wheat 5.8 2.8 2.6 33 3.5 0.894 0.745 3.9
7 Disc 1.2 mm Wheat 5.8 2.8 2.6 33 3.5 0.780 0.727 4.5
7 Disc 1.3 mm Wheat 5.8 2.8 2.6 33 3.5 1.033 0.819 3.4
7 Disc 1.4 mm Wheat 5.8 2.8 2.6 33 3.5 0.968 0.847 3.6
7 Disc 1.2 mm Sorghum 4.4 3.8 2.6 33 3.6 0.711 0.615 5.1
7 Disc 1.3 mm Sorghum 4.4 3.8 2.6 33 3.6 0.778 0.657 4.7
7 Disc 1.4 mm Sorghum 4.4 3.8 2.6 33 3.6 0.817 0.703 4.4
7 Disc 1.5 mm Sorghum 4.4 3.8 2.6 33 3.6 0.855 0.736 4.2
7 Disc 1.4 mm Barley 9.7 3.5 2.5 44 4.4 1.017 0.857 4.3
7 Disc 1.5 mm Barley 9.7 3.5 2.5 44 4.4 1.026 0.869 4.3
7 Disc 1.8 mm Maize 10.5 7.3 4.3 291 5.6 0.943 0.858 6.0
8 Disc ns Sorghum-wheat-soybean ng ng ng ng ng 0.659 0.559 ng
8 Disc ns Sorghum-barley-wheat ng ng ng ng ng 0.711 0.574 ng
8 Disc ns Sorghum-barley-wheat ng ng ng ng ng 0.808 0.664 ng
8 Disc ns Sorghum-barley-wheat ng ng ng ng ng 0.763 0.655 ng
8 Disc ns Wheat ng ng ng ng ng 0.706 0.536 ng
8 Disc ns Wheat ng ng ng ng ng 0.766 0.587 ng
8 Disc ns Wheat ng ng ng ng ng 0.923 0.750 ng
9 Hammer 3.6 mm Wheat 6.1 3.3 2.7 39 3.8 0.787 0.726 4.8
9 Hammer 4 mm Barley 8.5 3.4 2.5 39 4.2 0.906 0.764 4.6
9 Hammer 3.6 mm Sorghum 4.9 4.0 2.8 37 3.9 0.738 0.658 5.3
10 Roller ns Barley 8.2 3.3 2.3 34 4.0 2.446 1.235 1.6
10 Hammer ns Wheat ng ng ng ng ng 0.522 0.455 ng
10 Roller ns Sorghum ng ng ng ng ng 1.644 0.864 ng
18
The closer the discs were in the disc mills (reduced nip or gap), the finer were the milled
grains.
A 60% change in the size of the hammer mill sieve led to a 30% change in the average
particle size. However, for any hammer mill, there is a critical sieve size below which the
milling operation is not economic (cost and production of fines), irrespective of the small
particle size. This is applicable to disc- and roller-mills, and the need to reduce metal-to-
metal contact in both mills.
Mill settings varied with the grains.
Because of differences (type, variety and dimensions) in the grains being processed, it is
necessary to define a parameter that incorporates differences in dimensions, for example, of the
grains. Within a grain type, varieties that have big grains can fracture differently, and the
average particle size could be complemented by other fracturability indices in order to quantify
the milling operation. The dimensions and selected physical characteristics of the whole grains
are shown in Table 2. While grains are generally irregular, certain regular bodies can be used to
approximate their appearances (Sopade PA, Okonmah GN. (1993). The influence of
physicochemical properties of foods on their water absorption behaviour: A quantitative
approach. ASEAN Food Journal 8: 32). Sieving and other particle size analyses assume
spherical particles. Hence, if whole grains can be approximated to a sphere, the diameter of
such hypothetical sphere can be used to define the size reduction ratio (SRR) for comparison
purposes. The regular bodies that can be used to represent grains (cereals and pulses), and their
respective volumes are reproduced in Table 3.
Table 3: Approximate regular bodies for grains and the formulas for their volumes*
Grain Assumed regular shape Volume formula
Wheat Ellipsoidal 4
3 a
Barley Ellipsoidal 4
3 a
Sorghum Spherical 4
3
Maize Conical 2
3
Millet Conical 2
3
Soybean Ellipsoidal 4
3 a
Peanut Cylindrical
*a = major radius, b = minor radius and c = half of thickness
Using the above shapes, the size reduction ratio (SRR) for the hammer mills ranged from 3.3 –
6.4 (average = 5.1), for the disc mills, from 3.4 – 6.0 (average = 4.4) and for the roller mills,
from 1.6 – 5.9 (average = 3.1). From theory, the higher the SRR, the finer is the milling
operation. Hence, from all the fracturability indices, the roller mills generally milled to a
coarser size than the disc- and hammer-mills, with the latter generally yielding finer grains. It is
19
plausible, therefore, that hammer- and disc-mills will procedure better digested grains than
roller mills (see C.3.2). However, as discussed above, the thinly compressed grain particles
from roller mills, which might be retained on coarse sieves, can be equally digested as fine
particles when digestive enzymes saturate the whole area of the compressed particles.
Despite the importance of particle size in digestion, it was observed during the survey that most
mills did not have a procedure to regularly check the particle size of their milled grains prior to
use or packaging. Consequently, the mills did not know if the grains had been milled to an
acceptable size. This quality control issue can be addressed with the presence and use of a
simple sieving device. Although there is a commercial device, it was observed that its sieves
are of broad and non-specific sizes. There is a need for one device as simple as the existing
one, but cost effective, easy to use and with defined sieve sizes. Improving on the existing one,
a new design (not to scale) of a manual sieving device is shown in Figure 7.
Opening lock A, top B is gently slid right (as in the red arrow) towards the finest sieve before
placing solid body B to block the first (coarsest) sieve. Milled grains can then be added to just
fill the space between the solid body and top cover D. The solid body is then removed, and the
top is slid back left (as in the blue arrow) and locked (A). With the milled grains inside, the
device is held upright with the lock pointing up. Holding handle E for ergonometric reasons,
the device is shaken vertically and horizontally for about 5 min. The device can be placed
horizontally on two flat supports to read the volume of the milled grains retained on each sieve.
The volumes are totalled to evaluate the goodness of the milling operation, with good milling
defined as leaving less than 20% of the total volume retained on sieves larger than 1 mm
aperture or sieve size.
C.3 Laboratory analysis of selected commercially milled
grains
C.3.1 Evaluation of the in-vitro protein digestion procedure Prior to studying the in-vitro protein digestion of the samples in the present study, some feed
samples and ingredients, cereals and pulses, were studied for their in-vitro protein digestion.
The samples were milled to pass through a 1-mm. sieve. The protein digestograms of the
selected cereal and pulses from these samples are shown in Figure 8 with the pH reducing with
digestion time as amino acids were produced. Remarkably, the digestograms were suitably
described (r2 > 0.9; p < 0.001) by the modified first order kinetic model (Eq. (4)). The
deviation of the experimental data from the predicted data during the early part of the
digestogram could be due to experimental errors or a possible biphasic digestion process,
which is complex to model, but it is being investigated. Table 4 shows the protein digestion
parameters for some of the samples, with and without treatments with a protease (250 ppm,
Avizyme 1510 Protease, 40,000 units/g, Genencor International, Netherlands).
The suitability of the procedure was further investigated with various samples including mash
and pelleted samples from wheat, barley and sorghum grains, which were hammer-milled with
different sieves (2.0 and 3.2 mm) prior to pelletisation. The protein digestograms (Fig. 9) are
similar to those reported before (e.g. Fig. 8), sensitive to processing and materials, and the
modified first-order kinetic model was also suitable in describing the digestograms. Table 5
summarises the protein digestion parameters for this class of samples.
20
Figure 7: Design of a manual sieving device
2.50 aperture, 0.71 diameter 304S/S
1.40 aperture, 0.71 diameter 304S/S
1.00 aperture, 0.56 diameter 304S/S
0.71 aperture, 0.56 diameter 304S/S
0.50 aperture, 0.32 diameter 304S/S
0.10 aperture, 0.071 diameter 304S/S
www.locker.com.au
90 mm 60 mm 50 mm
100 mm
450 mm
20 20 20 20 20 20
60 60 60 60 60 60
A
C
E
B
D
21
Table 4. Effect of protease treatment on in-vitro protein digestibility of feed
ingredients*
Sample Protein content IVPD (%) Kpr x 10-3
(min-1
) (g/100 g solids) Protease
No
Protease
Protease
No
Protease
Canola meal 42.3c 92.8abcd 92.4abcd 329a 260a
Meat meal 54.8b 93.4abcd 91.1abcd 436a 288a
Millrun 22.5f 90.8abcd 89.8abcd 341a 293a
Mung beans 29.3e 90.7abcd 86.6cd 400a 254a
Red sorghum 14.5g 87.5abcd 84.8d 324a 242a
Soyabean 55.6b 94.1abc 94.4abc 289a 302a
Sunflower 37.0d 89.6abcd 87.2bcd 321a 278a
White sorghum 12.9g 87.5abcd 85.0d 403a 329a
Mean of the 15 samples
studied Not applicable 91.9A 90.4B 342A 297B
*For protein, and between protease and no protease, for IVPD, K and mean, values with the same letters are non-
significant (P>0.05).
Table 5: Effect of processing and sample diversity on in-vitro protein digestibility
Sample
Sieve
(mm)
State`
Protein
(g/100g solids)
Kpr x 10-3
(min-1
)
IVPD
(%)
Barley 2.0 Mash 12.8ef 457.8ab 92.8ab
Barley 2.0 Pellet 12.6f 443.3ab 99.9a
Barley 3.2 Mash 11.9ghi 357.1bcd 88.9b
Barley 3.2 Pellet 12.3fg 457.9ab 93.0ab
Canola meal na† na 44.0c 153.0f 89.0b
Commercial starch na na 0.6k 268.1cdef 78.1c
Cooked Rice na na 10.4j 286.3cdef 93.5ab
Hydrolysed casein na na 95.4a 536.0a 94.0ab
Sorghum 2.0 Mash 11.4i 387.6bc 86.9b
Sorghum 2.0 Pellet 11.7hi 203.4ef 90.1b
Sorghum 3.2 Mash 12.0gh 208.3ef 86.2bc
Sorghum 3.2 Pellet 11.8ghi 362.9bcd 89.9b
Soyabean meal na na 54.0b 241.3def 89.8b
Uncooked Rice na na 9.8j 542.3a 91.9ab
Wheat 2.0 Mash 14.6d 377.9bc 94.5ab
Wheat 2.0 Pellet 14.4d 347.1bcd 93.5ab
Wheat 3.2 Mash 13.2e 360.5bcd 91.0b
Wheat 3.2 Pellet 14.2d 336.9bcde 92.1ab *Values with the same letters are non-significant (p>0.05) †na = not applicable
22
Figure 8: Protein digestograms of feed ingredients
6.6
6.8
7.0
7.2
7.4
7.6
7.8
8.0
0 2 4 6 8 10 12
Experimental
Predicted
Canola meal
6.6
6.8
7.0
7.2
7.4
7.6
7.8
8.0
0 2 4 6 8 10 12
Experimental
Soybean meal
6.6
6.8
7.0
7.2
7.4
7.6
7.8
8.0
0 2 4 6 8 10 12
Experimental
Lupins
6.6
6.8
7.0
7.2
7.4
7.6
7.8
8.0
0 2 4 6 8 10 12
Experimental
Field peas
6.6
6.8
7.0
7.2
7.4
7.6
7.8
8.0
0 2 4 6 8 10 12
Experimental
Mung beans
6.6
6.8
7.0
7.2
7.4
7.6
7.8
8.0
0 2 4 6 8 10 12
Experimental
White sorghum
pH
Time (min.)
23
Figure 9: Protein digestograms of processed and non-processed diverse materials
6.6
6.8
7.0
7.2
7.4
7.6
7.8
8.0
0 2 4 6 8 10 12
Wheat 2 mm-sieve Mash
Experimental
6.6
6.8
7.0
7.2
7.4
7.6
7.8
8.0
0 2 4 6 8 10 12
Barley 3.2 mm-sieve Pellet
Experimental
Predicted
6.6
6.8
7.0
7.2
7.4
7.6
7.8
8.0
0 2 4 6 8 10 12
Commercial starch
Experimental
6.6
6.8
7.0
7.2
7.4
7.6
7.8
8.0
0 2 4 6 8 10 12
Sorghum 3.2 mm-sieve Pellet
Experimental
6.6
6.8
7.0
7.2
7.4
7.6
7.8
8.0
0 2 4 6 8 10 12
Rice
Experimental
6.6
6.8
7.0
7.2
7.4
7.6
7.8
8.0
0 2 4 6 8 10 12
Cooked rice
Experimental
pH
Time (min.)
24
Hence, from the various in-vitro protein digestion methods investigated, the adapted three-
enzyme procedure is the simplest and the most reproducible, can show differences between
samples, and it is sensitive to processing, protein content as well as particle size of samples.
Studies on black-eye beans (or cowpea), a pulse, revealed, as obtained for starch digestion, the
rate of protein digestion was inversely related to the square of the particle size of the cryo-
milled beans, but the hammer-milled beans did not show the same relationship (Tinus T,
Damour M, van Riel V, Sopade PA. (2012). Particle size-digestibility relationships, if starch,
does protein? Journal of Food Engineering (submitted)).
C.3.2 Laboratory analysis of commercially milled grains As indicated above (B.2), the disc-milled sorghum and wheat from mill 7 (Table 2) were
analysed for their in-vitro starch and protein digestion. All the samples essentially exhibited
monophasic digestograms (Fig. 10), which were suitably (r2 > 0.9; p < 0.001) described by the
modified first-order kinetic models (Eq. (4) and Eq. (5)). The parameters of the models, and
properties of the samples are summarised in Table 6. It is worth noting that pending detailed
statistical analysis, further discussions are based on the mean values presented in Table 6.
The sorghum had a slightly higher moisture content, but both grains were within the safe
storage moisture content (14 – 15%) for grains. The wheat had a higher protein content but a
lower starch content than the sorghum. The differences within a grain were most likely
experimental errors assuming that the samples commercially-milled in each grain were
homogenous. The proteins in the samples were more rapidly digested (75X) than the starch.
The higher rate of protein digestion than starch was observed with other heat-processed and
non-processed samples. With three enzymes in each procedure, and possibly at excess
concentrations, this could indicate that proteins are easier to digest than starch and would be
valuable in understanding asynchrony of nutrients in feeds.
Even though there were grain differences, protein or starch digestibility of the grains was not
materially different from one another. Sorghum is generally less digestible than other cereals,
and the results obtained in this study could be because commercial grains can be mixtures of
varieties with possibly uncontrolled G x E effects. Although the extent to which commercial
situations could influence the well-known trends is unknown, the present results show that
commercial samples need to be carefully sampled and characterised. With commercial samples
representing field situations, trends (e.g. particle size-digestibility) obtained with laboratory
samples need to be re-evaluated with commercial samples. Contrary to previous studies, the
rates of protein and starch digestion did not exhibit any relationship with the square of particle
size of either the milled sorghum or wheat (Fig. 11). There are no immediate reasons for this
observation, but it might be related to the non-homogenous nature of commercial samples, as
highlighted above, and follow-up studies are planned in this area with more commercial
samples.
25
Figure 10: Protein and starch digestograms of the commercially-milled sorghum and wheat
7.0
7.2
7.4
7.6
7.8
8.0
0 2 4 6 8 10 12
Wheat 1.2 mm
Experimental
Predicted
7.0
7.2
7.4
7.6
7.8
8.0
0 2 4 6 8 10 12
Sorghum 1.3 mm
Experimental
0
10
20
30
40
50
60
0 40 80 120 160 200 240
Wheat 1.1 mm
Experimental
0
10
20
30
40
50
60
0 40 80 120 160 200 240
Sorghum 1.4 mm
Experimental
pHD
iges
ted
star
ch (
g/10
0gdr
y st
arch
)
Time (min.)
26
Table 6: Digestion and chemical properties of the commercially-milled sorghum and wheat
Sample Moisture
Content
(g/100g
sample)
Protein
content
(g/100g
solids)
Starch
content
(g/100g
solids)
In-vitro digestion
Protein Starch
Kpr x 10-3
(min-1
)
IVPD
(g/100g
protein)
Do
(g/100g
dry starch)
D∞
(g/100g
dry starch)
Kst x 10-3
(min-1
)
AUCx103
(g.min/100g
dry starch)
PreD240
(g/100g
dry starch)
Sorghum_1.2 15.4±0.37 9.0±0.27 50.3±3.24 238±0.4 77.9±2.94 0.0±0.00 100.0±0.00 3.4±0.25 7.6±0.43 56.0±2.67
Sorghum_1.3 14.6±0.01 9.9±0.25 60.7±2.61 195±1.9 82.0±1.20 0.0±0.00 100.0±0.00 2.6±0.02 6.2±0.04 46.8±0.28
Sorghum_1.4 15.5±0.03 10.0±0.32 56.0±1.93 221±0.5 77.9±2.63 0.0±0.00 100.0±0.00 2.7±0.16 6.2±0.30 47.1±1.98
Sorghum_1.5 15.3±0.16 9.9±0.37 50.7±1.23 320±2.4 83.1±3.69 0.0±0.00 100.0±0.00 3.0±0.05 6.8±0.09 50.9±0.61
Wheat_1.1 14.2±0.21 13.9±1.13 45.9±2.46 263±1.7 85.9±0.89 0.0±0.00 89.9±10.51 3.3±1.14 6.6±0.99 48.2±5.38
Wheat_1.2 14.1±0.00 14.8±0.74 48.7±3.21 236±2.4 84.0±2.94 1.0±1.40 75.6±14.97 3.7±0.38 6.3±0.54 44.7±5.50
Wheat_1.3 13.7±0.02 15.2±0.30 43.6±1.6 241±1.9 87.3±5.37 0.9±1.21 78.6±8.01 3.8±0.17 6.6±0.68 47.5±5.59
Wheat_1.4 14.2±0.00 15.4±0.31 46.3±0.8 218±1.4 88.1±3.59 0.7±0.95 68.8±8.31 3.3±0.22 5.2±0.71 37.7±5.70
27
Figure 11: Plots of rates of starch and protein digestion, and square of geometric mean particle size
15
16
17
18
19
0.006 0.007 0.008 0.009 0.010 0.011
Wheat - starch
17
18
19
20
21
22
23
24
0.005 0.006 0.007 0.008
Sorghum - starch
220
230
240
250
260
270
280
0.006 0.007 0.008 0.009 0.010 0.011
Wheat - protein
180
200
220
240
260
280
300
320
0.005 0.006 0.007 0.008
Sorghum - protein
1/K
pr,
1/K
stx
103
(sec
)
Dgw2 (cm)
28
The differences in the particle size distributions of the mills were discussed in C.2, and their
implications for digestibility of diets. Although a detailed study is proposed in follow-up
studies, preliminary studies on the in-vitro starch digestion of commercially milled sorghum
revealed hammer milling could yield better digested diets than roller milling (Fig. 12A). Wheat
was also milled in laboratory Buhler-, disc- and hammer-mills, and Figure 12B shows that
while both disc- and hammer-mills could produce about the same digestibility, Buhler-mill
would yield a better digested diet. In comparing mills, however, it should be stressed that mill
settings can be different, and these need to be stated for ease of understanding, scale-up and
transfer to commercial operations for maximum digestibility of pig feeds.
D. Conclusions There were three types of mills mostly used in the Australian feed industry, with hammer mills
being the most prominent. There appeared to be a shift to disc mills with a prime motive to
increase digestibility of diets through the manipulation of particle size distributions. Although
mill type can be seen to mostly impact mash diets, because particle size controls heat and mass
transfers, particle size is equally important in pelleted diets. This calls for a careful control of
particle size distributions, which define particle sizes of milled grains. The present study
revealed that broad to narrow distributions were obtainable from the feed mills, with
differences in the proportions of large particles (> 1 mm). Large particles in mash or pelleted
diets reduce digestible energy, and might pass through the digestive tract undigested. Large
particles were observed in ileal digesta, even though their proportions in the digesta were less
than those in the diets. The present study identified the need for a simple, manual, robust and
cost effective sieving device for quality control of feed milling operations. Such device had
been designed, and will be fabricated and field-tested in follow-up studies in this project.
Furthermore, an in-vitro protein digestion procedure was tested and found to be sensitive to
analytes and processing. It is a rapid (about 15 min. analysis) method, and it could be a
component of a quality control strategy in feed mills to complement the in-vitro starch
digestion procedure reported earlier. Both methods will help to understand how milling
influences digestion, as a first step, in order to optimise mill settings for maximum digestion.
However, using both in-vitro methods, no relationship was obtained between the reciprocal of
the rates of digestion of the commercially-milled sorghum and wheat investigated, and the
square of their particle sizes. While the particle size-digestibility relationship for starch has
been proven for other samples, the absence of such for either starch or protein in the present
study raises questions about sampling of commercial materials, which can be mixtures of grain
varieties with possibly no documented G x E effects. However, commercial samples represent
field situations. This notwithstanding, the present study raises issues for follow-up studies vis-
a-vis type of mill, mill setting and on-site particle size analysis to evaluate milling operations
for optimum energy delivery from pig feeds. The present study also calls for continued
collaborations with commercial feed mills so that outcomes of laboratory and animals studies
can be disseminated and used to improve milling of grains (cereals and pulses) in the
Australian feed industry.
29
Figure 12: Starch digestograms showing differences between mills
0
10
20
30
40
50
60
0 40 80 120 160 200 240
Sorghum - Feed mill 1
Hammer mill
Roller mill
Predicted
A
0
10
20
30
40
50
0 40 80 120 160 200 240
Wheat
Buhler mill
Disc mill
Hammer mill
Predicted
B
Dig
est
ed
sta
rch
(g/1
00g
dry s
tarch
)
Time (min.)
30
E. Further research needs and recommendations The following have been identified for future research needs:
Evaluation of particle size distributions at different settings in a commercial mill. This
will involve feed mills will hammer-, disc- and roller-mills.
Identification of mill settings required to produce narrow and broad distributions, and
either, at a known average particle size.
Fabricate and supply manual sieving device to selected feed mills in the first instance.
Animal trials (feeding and metabolism) to examine the concept of particle size
distributions.
Laboratory studies on how particle size affects protein digestion.
Examination of the relationships between particle size and protein digestion in an
expanded study.
Acknowledgements
The authors gratefully acknowledge the contributions towards outcomes from this project by
Ms Sara Willis of the Department of Employment, Economic Development and Innovation
(DEEDI), Toowoomba, Qld 4350, and Ms Jing Zhang and Ms Su Sin Koa of the Centre for
Nutrition & Food Sciences, University of Queensland, St Lucia, QLD 4072, as well as the
managements of the feed mills that participated in the project.
