Copyright © 2012 American Scientific Publishers Journal of

All rights reserved Biobased Materials and Bioenergy

Printed in the United States of America Vol. 6, 431–439, 2012

Fungal Mycelium and Cotton Plant Materials in the

Manufacture of Biodegradable Molded Packaging
Material: Evaluation Study of Select Blends of
Cotton Byproducts
G. A. Holt , G. McIntyre2, D. Flagg2, E. Bayer2, J. D. Wanjura1, and M. G. Pelletier1

RESEARCHARTICLE
1
USDA-ARS Cotton Production and Processing Research Unit 1604 E. FM 1294, Lubbock, TX 79403, USA
2
Ecovative Design, LLC, 60 Cohoes Avenue, Green Island, NY 12183, USA

Since polystyrene is non-biodegradable,Delivered by Ingenta to:a biodegradable material that is

eco-friendly is being sought as a substitute for packaging and insulation Greg Holtboard consumers. One
such process, developed by Ecovative Design, LLC, involves growing IP : 199.133.133.66fungal species
on agricultural biomass to produce an ecofriendly packaging product (EcoCradle™ Thu, 29 Nov 2012
23:20:57) and insulation panels (Greensulate™). The objective of this research was to develop and
evaluate six blends of processed cotton plant biomass ( CPB ) materials as a substrate for colonization of
selected fungi in the manufacture of molded packaging material. The blends were comprised of processed
CPB, cotton seed hulls, starch, and gypsum. The four ingredients were the same mix percentage for all six
blends with the particle size of the CPM being the only difference. CPB particles sizes ranged from 0.1 to 51
mm. Tests were conducted to evaluate the physical and mechanical properties of the six CPB blends. Test
results revealed blends that met or exceeded like characteristics of extruded polystyrene foam.

Keywords: Composite, Cotton, Mycelium, Biobased, Biodegradable.

1. INTRODUCTION studied and used to manufacture environmentally friendly

Polystyrene is a hydrocarbon based material typically used
in the manufacture of packaging materials. Marketed
under the name Styrofoam™, this lightweight material is
hydrophobic, resistant to photolysis, and is not subject to
decomposition or decay.1 These characteristics are
attractive to shippers and the packaging industry, but they
create problems with respect to recycling, reuse, and
landfill operation.2 Recent scientific investigation resulted
in development of an economically viable,
environmentally friendly replacement for polystyrene
packaging materials. The packaging material evaluated in
our study is a composite containing selected agricultural
residues and a specific fungus.
Annually renewable crops and their agricultural
products.7–10 The combination of agricultural residues and
fungi have been evaluated for fungal cultivation11–15 and
improving bonding properties of agricultural fibers in the
manufacture of composites.1617 However, a method for
producing a composite comprised of agricultural residues
and fungi was not found in the literature.
Bayer et al.18 and Bayer and McIntyre19 developed
processes that involve growing fungal species on
agricultural residues, such as cotton plant material, to
produce an environmentally-friendly packaging material.
The objective of the present study was to develop six
blends of specifically processed cotton plant material for
use with the Bayer et al.18 and Bayer and McIntyre19
processes. Each of the six blends was used to produce a
packaging material that was subjected to standard test

431
 Fungal Mycelium and Cotton Plant Materials in the Manufacture of Biodegradable Molded Packaging Material Holt et al.

residues have been researched extensively as materials
that could potentially enhance the properties of composite
products made from fossil fuel based derivatives that are
more resistant to biodegradation. 3–6 In addition to the crops
and agricultural residues, fungi and/or their constituents
have been
∗
Author to whom correspondence should be addressed.
Email: greg.holt@ars.usda.gov
methods for compressive strength,20 flexural strength,21
modulus of elasticity,21 density,22 dimensional stability,22
accelerated aging,23 water absorption,24 cone calorimetry,25
and thermal conductivity.26 The cotton plant material used
in this study was a byproduct of typical mechanical
harvesting and ginning practices in the United States
which generate approximately 2.5 Mg of cotton
byproducts across the U.S. cotton belt each year.27

J. Biobased Mater. Bioenergy 2012, Vol. 6, No. 4 1556-6560/2012/6/431/009 doi:10.1166/jbmb.2012.1241

2. MATERIALS AND METHODS

2.1. Cotton Plant Biomass and Blends

The cotton plant material was obtained from a cotton gins

agricultural waste stream. Specifically, the cotton carpel
(bur) was obtained from the extractors 27 (HỆ THỐNG
TRÍCH LY) in the United States Department of
Agriculture–Agricultural Research Service (USDA-ARS)
gin lab in Lubbock, Texas. The extractors removed
approximately 55% of the total waste stream, primarily
sticks and carpel, from the incoming seedcotton before Fig. 1. Some of the cotton plant material used in this study before ( left )
the seedcotton entered the gin stand(s). The cotton carpels and after (right) processing. MỘT SỐ NGUYÊN LIỆU TRƯỚC VÀ
were processed through a Jacobson model 4D hammer- SAU SỬ LÝ
mill (Jacobson Machine Works, Minneapolis, MN) or a
Reynolds Engineering model T00-18 attrition mill
material. Once all the ingredients were added, the blend
(Reynolds Engineering and Equipment, Muscatine, IA)
was allowed to mix for 7 min, at maximum agitation (15
and then sorted across a Triple/S Dynamics model
rpm). Upon completion of the mixing, the blend was
VCSUF-24X12-3 vibratory conveyor (Triple/S
emptied into a tote bag, labeled, and stored until
RESEARCHARTICLE

Dynamics, Inc., Dallas, TX) to obtain the specific particle

cotton plant material particles were sized to be within the
desired range, 0.1 to 51 mm.
Blending of the cotton plant material was accomplished
with a Davis 1.4 Mg ribbon mixer (H.C. Davis Sons
Manufacturing Co., Inc., Bonner Springs, KS). Each
blend was processed in 159 kg batches. The constituent
materials used in the six blends evaluated were comprised
of:
(1) processed cotton
carpel,
(2) cotton seed hull,
(3) starch, and
(4) gypsum.
The primary ingredient in each of the six blends was the
processed cotton carpel. For all six blends, the ingredients
were added at the same percentages with the only
difference being the particle size range of the processed
cotton carpel (Table I). Each ingredient was added one at
a time into the mixer and agitated while adding between
1.5 to 2.5 l of water, to minimize dust and promote
adhesion of the starches and gypsum to the cotton plant
Table I. Particle size ranges for cotton plant material used in each of the
six blends evaluated in this study.
Primary particle size range of
Blends a
processed cotton plant material (mm)
1 28 – 51
2 12 – 28
3 0.1–12
4 12–51
5 0.1–12, 28–51
6 0.1–51
Notes: aPrimary ingredient of each blend was the processed cotton plant material.
The other ingredients in each blend were starch, gypsum, and cottonseed hulls.
2.2. Composite Fabrication

size ranges needed for each blend (Table I). Delivered by Ingenta to:The cotton shipment to Ecovative Design’s
facility in Green Island, carpel material, for one of the blends, before and after Greg HoltNY. Specifics related to
constituent fraction of the blends processing is shown in Figure 1. 432 material IP
The sizedJ. Biobased : 199.133.133.66
Mater. Bioenergy 6, 431–439,
was2012
are
considered proprietary information by Ecovative packaged and stored in a dry location until Thu, 29 Nov 2012
23:20:57blending. The Design, LLC.
Holt et al.Fungal Mycelium and Cotton Plant Materials in the Manufacture of Biodegradable Molded Packaging Material

At Ecovative’s research laboratory in Green Island, New pasteurizer and cooler speeds, and specific processes
York, is the pilot manufacturing plant. A schematic of the applied not listed are considered proprietary information
pilot plant process is shown in Figure 2. At the pilot plant, by Ecovative Design, LLC.

RESEARCHARTICLE
Delivered by Ingenta to:
Greg Holt
IP : 199.133.133.66
Thu, 29 Nov 2012 23:20:57

Fig. 2. Schematic of Ecovative’s pilot manufacturing facility used to produce the cotton plant and fungal mycelium based molded packaging specimens
the tote bags with each blend were emptied into a bulk bin
with a live-bottom auger that fed a pasteurizer where the
material was sterilized at 115 C for approximately 28 min.
Exiting the pasteurizer, the blend was gravity fed into a
water-jacketed auger (cooler) where it was cooled below
35 C. Upon exiting the cooler, the blend was inoculated
with the fungus, Ganoderma sp., using a specified grain or
liquid substrate as the carrier. After inoculation, the blend
was discharged into a plastic mold, referred to as a tool,
which was in the desired shape of the piece to be
fabricated. The material was gently hand-packed in the
tool and any excess was removed, the tool was then
sealed in order to maintain a consistent micro-
environment for fungal propagation (Fig. 3). The filled
tool was incubated on a bread rack at 21 C for 5-days at
which time the fungal mycelium colonized the blend.
Figure 4 shows the fungal colonization of one of the
blends over a three day period. After 5 days, the part was
removed from the tool and placed in 60 C convection
oven for 8 h, which inactivated the fungus and prevented
reanimation. After drying, the pieces were stored at
ambient laboratory conditions (approximately 21 C and
30% RH) until testing. Figure 5 shows a typical part after
drying. Specifics related to quantity of inoculum,
evaluated in this study.
2.3. Analytical Testing
Each of the six blends was used to produce a packaging
material that was subjected to standard test methods for
compressive strength,20 flexural strength21 (Fig. 6),
modulus of elasticity,21 density,22 dimensional stability,22
accelerated aging,23 water absorption,24 cone calorimetry25
(Fig. 7), and thermal conductivity.26 Cone calorimetry
(flame retardance characteristics) was performed at the
Worcester Polytechnic Institute (WPI) Fire Research
Laboratory in Worcester, MA. Specimens were tested in
horizontal orientation at 50 kW/m2 heat flux. All other
analyses were conducted at the Ecovative research
laboratory. We did not evaluate expanded polystyrene
samples in this study. Numerous sources of information
pertaining to physical and mechanical properties of
expanded polystyrene are available in the public domain.28
– 31
2.4. Data Analysis

J. Biobased Mater. Bioenergy 6, 431–439, 2012 433

 Fungal Mycelium and Cotton Plant Materials in the Manufacture of Biodegradable Molded Packaging Material Holt et al.

Two types of inoculum (grain and liquid substrate) were

applied to each of six cotton plant material blends for a

434 J. Biobased Mater. Bioenergy 6, 431–439, 2012

Holt et al.Fungal Mycelium and Cotton Plant Materials in the Manufacture of Biodegradable Molded Packaging Material

Fig. 3.

Selecting the lid for the tool containing inoculated cotton plant material substrate (left)
and snapping the lid in place (right) to maintain micro-environment for optimum growth.
Fig. 5. Finished fungal mycelium molded packaging piece after drying.

total of 12 treatments. Each treatment was replicated from 3 to 12 times

depending on the test method and property being evaluated.
Standard analysis of variance techniques were used to analyze the
data to determine statistically
Delivered by
Ingenta
to:significant
differences among
the 12 treatments by
RESEARCHARTICLE

the
Greg HoltTukey-Kramer Honestly Significant Difference (HSD) test
IP :
199.133.133.66(rele
ase 9.2, SAS Institute
Inc., Cary, NC) at the
95% conThu, 29 Nov
2012
23:20:57fidence level.

Fig. 4. Time and fungal colonization over a 3-day period. The white specks are the
living fungus.
sequence showing the inoculated cotton plant material substrate (day 0) Fig. 6. Flexural strength testing of one of the cotton plant material
test specimens.

J. Biobased Mater. Bioenergy 6, 431–439, 2012 435

 Fungal Mycelium and Cotton Plant Materials in the Manufacture of Biodegradable Molded Packaging Material Holt et al.

liquidbased inoculum) exhibited the greatest measured

surface area contraction (2.4%). Grain 3, 4, 5, and Liquid
6 were similar in the percent contraction and significantly
lower than Grain 2, Liquid 1, 2, 3, 4, and 5.
The importance of dimensional stability is related to
tool design. The larger the percent contraction the more
oversized the tool needs to be for the finished product to
be within desired specifications. Another factor
influencdue to the greater mass of the grain-based
inoculum versus the liquid-based inoculum. The density
adjusted values for FS show Grain 1 and Grain 6 with the
highest values at 26 kPa and Liquid 2 with the lowest at 7
Fig. 7. Cone calorimeter test samples of two of the treatments evaluated
kPa. The EM was significantly higher for Liquid 5 (674
in this study, Grain 6 (left) and Grain 4 (right), after testing. kPa) than all other treatments with Liquid 2 having the
lowest, 123 kPa. Compressive strength was significantly
3. RESULTS AND DISCUSSION
higher for Liquid 3 (72 kPa) than for all other treatments
3.1. Physical Properties with Liquid 4 having the lowest CS at 1.1 kPa.
Sample degradation associated with FS, EM, and CS
Figure 8 shows a comparison of the dimensional stability resulting from accelerated aging testing is shown in the
(surface area contraction) of the treatments after drying. second column of Table II. The percent degradation data
Blends inoculated with the grain-based substrate showed was calculated according to the equation in the standard:23
less surface area contraction than did blends inoculated
with the liquid-based substrate. Grain 3 (Blend 3, Degradation Percentage
grainbased inoculum) had the smallest measured surface
= (Conditioned test value/as received test value)∗100
area contraction (0.64%), whereas Liquid 5 (Blend 5,
ing tool design is contraction variability. The Delivered by Ingenta to:more vari- Therefore, values closer to the base
line (100%) exhibit able a blend/inoculum combination is, the more difficult Greg Holtless degradation than samples
with values further from the it is to produce parts that are consistently within IP : 199.133.133.66dimen- baseline. FS
degradation for Grain 5, 1, 6, and Liquid 3 sional tolerances of customer specifications. All Thu, 29 Nov 2012
23:20:57treatments exhibited little degradation from aging. Liquid 5’s FS was
had similar standard mean errors associate with percent reduced almost in half as a result of aging whereas the FS
contraction (0.093 to 0.108), so the means are a reliable of Grain 3 and Liquid 2 exhibited increased stiffness due
indicator of the contraction expected when designing tools to aging. EM for Grain 6 had the largest change in percent
for a given treatment. degradation of 318.6%. Liquid 5 had the largest reduction
The flexure strength (FS), elastic modulus (EM), and in EM at 43.6%. The treatments that had the largest
compressive strength (CS) in Table II are normalized to a percent CS degradation were Liquid 5 (250.1%) and
standard density of 32.04 kg/m3 since this is the density of Liquid 3 (6.8%). The CS degradation was least for Grain 4
the polystyrene packaging the EcoCradle material can (92%) and Grain 1 (110%). Overall, Grain 1 had the most
replace in the market. The density of the treatments ranged consistent performance, by exhibiting some of the lowest
from 66.5 kg/m3 to 224 kg/m3. The density for grain degradation values for FS, EM, and CS compared to all
treatments was higher than for the liquid treatments other treatments.
RESEARCHARTICLE

436 J. Biobased Mater. Bioenergy 6, 431–439, 2012

 Holt et al.Fungal Mycelium and Cotton Plant Materials in the Manufacture of Biodegradable Molded Packaging Material

Fig. 8. Average surface area contraction (%) or shrinkage of the sample pieces made from each treatment after oven drying. Bars with the same letters
inoculum methods.
Physical properties (density adjusted)
a Water absorption
Response variable Flexure strength Flexure strength degradation
are not significantly different at the 0.05 level of significance.
Table II. Physical properties from molded packaging material produced Table III. Water absorption testing results from molded packaging from six
cotton-based substrate blends and two fungal inoculum methods. material produced from six cotton-based substrate blends and two fungal
Standardb ASTM C203 ASTM C481 Response Gain after Gain after Gain after

Units (kPa ) (% ) variable 75 hr 3 hr 168 hr

c c Standard ASTM C1134 ASTM C1134 ASTM C1134
Inoculum/blend Value Inoculum/blend Value
Units (% ) (% ) (% )
Liquid
Grain 11 208A
26 1AB C Liquid
Liquid 62 1557A
196 7AB Liquid 1 207A B Liquid 4 392A B Liquid 4 1780 A
Inoculum/
Grain 5 Inoculum/ Inoculum/
Liquid
Grain 65 196A
25 9AB C Liquid
Grain 13 1151A
186 0AB 171B Cb Liquid 1 342A Bb Liquid 1 1688 Ab
blend Value blend Value blend Value
Grain 23
Liquid 23 5ABBC
164A Grain 24
Grain 181 2ABB
1116A Liquid 2 139B C Grain 5 298A B Liquid 5 1637 A
Grain 1 30 7A Grain 1 48 0A Grain 1 198 1A
Grain 5 219A B C Liquid 4 1583A B

Liquid 5 115B C Liquid 5 258A B Grain 5 1550 A

Grain 3 123B C Grain 6 1005A B
Liquid 4 112B C Liquid 2 252A B Liquid 2 1536 A
Grain 4 104B C Grain 1 1003A B
Liquid 3 103B C Liquid 6 214A B Liquid 6 1324 A
RESEARCHARTICLE

Liquid 6 101B C Grain 5 1001A B 9B C Grain 3 179B Grain

2 1269 A Grain 2 9
Liquid 4 97B C Liquid 3 950A B Grain 3 93B C Grain 2 177B Grain 4 1103 A
Liquid 2 70C Liquid 5 600B Grain 4 90B C Liquid 3 155B Liquid 3 1061 A a Elastic modulus Elastic
modulusDelivered by Ingenta to: degradation Liquid 6 70C Grain 4 122B Grain 6 943 A
Response variable

Standardb ASTM C203 ASTM C481 Greg HoltGrain 6 64C Grain 6 108B Grain 3 935 A
)
Units (kPa) (% IP : 199.133.133.66Notes: ASTM = American Society for Testing and Materials. Means within the
a b

Inoculum/blend Value c Inoculum/blendThu, 29 Nov 2012 23:20:57Value c same column followed by different letters in the corresponding row are
statistically
different at the 0.05 level of significance.

Liquid 5 6745A Grain 6 3186 A Grain 2 3956B Liquid 2 1894A B

3.2. Water Absorption
Grain 1 3681B C Grain 3 1659A B

Liquid 4 3318B C Grain 4 1632A B The water absorption results (Table III) show the percent
Grain 3 3174B C Liquid 6 1507A B weight gain after 0.75 h, 3 h, and 168 h. After 0.75 h,

J. Biobased Mater. Bioenergy 6, 431–439, 2012 437

 Fungal Mycelium and Cotton Plant Materials in the Manufacture of Biodegradable Molded Packaging Material Holt et al.

Grain 6 (6.4%) and Liquid 6 (7.0%) had the lowest water

Liquid 6 3088B C Liquid 3 1421B
absorption whereas Grain 1 (30.7%) and Liquid 1 (20.7%)
Grain 6 2884B C Grain 2 1347A B had the highest. Grain 1 had significantly higher water
Grain 5 2834B C Liquid 4 1219B absorption after 0.75 h than all other treatments except
Liquid 1 2538B C Grain 1 984B Liquid 1.
Grain 4 1991B C Liquid 1 955 B
Liquid 2 1228C Liquid 5 436 B
Table IV. Conductivity and R-values from molded packaging material
Liquid 3 3442B C Grain 5 1641 B

a
producedfrom six cotton-basedsubstrateblends and two fungal inoculum
variable strength degradation methods.
b
Standard ASTM C165 ASTM C481
Units (kPa) (%) Thermal properties
c c
Inoculum/blend Value Inoculum/blend Value Response variable R -value Conductivity
a
Standard TPS500 TPS500
Liquid 3 72 2A Liquid 5 250 1A Units – W/m∗K
Liquid 1 33 5B Grain 3 169 5A B b b
Inoculum/blend Value Inoculum/blend Value
Grain 6 11 0B C Grain 2 139 9A B
Grain 2 9 0B C Liquid 4 120 2A B Liquid 6 1 51A Grain 3 0 18A

Response Compressive Compressive strength

Grain 6 100A B Grain 2 013A B
Grain 1 096B Grain 4 013A B
Grain 5 85B C Grain 1 1101A B Liquid 4 087B Liquid 1 012A B
Grain 4 62C Grain 4 917A B Liquid 5 084B Liquid 3 012A B
Grain 1 54C Liquid 6 877A B Grain 3 082B Liquid 6 010 B
Liquid 2 33C Grain 6 605A B
Liquid 6 30C Liquid 2 477A B Notes: aTSP = Transient plane source from therm test incorporated, Frederiction,
Grain 3 23C Grain 5 421A B Nebraska, USA. bMeans within the same column followed by different letters in the
Liquid 5 14C Liquid 1 81A B corresponding row are statistically different at the 0.05 level of significance.
Liquid 4 11C Liquid 3 68B After 3 h, Grain 1 (48.0%) still had the highest water
absorption and Grain 6 (10.8%) the lowest. However,
Notes: aResponse variables are normalized to a density of 32.04 kg/m 3. Degradation Grain 1 was only significantly higher than five other
= (Tested specimen value/as received specimen value)∗100. Degradation values of treatments: Grain 2, Grain 3, Grain 4, Grain 6, and Liquid
100 indicate no change before and after testing. bASTM = American Society for
3.
Testing and Materials. cMeans within the same column followed by different letters
in the corresponding row are statistically different at the 0.05 level of significance.
Liquid 3 140A B Liquid 4 017 A
Liquid 1 128A B Grain 1 016A B
Grain 2 115A B Liquid 5 015A B
Grain 4 114A B Grain 6 015A B
Grain 5 112A B Liquid 2 015A B
Liquid 2 105A B Grain 5 013A B

438 J. Biobased Mater. Bioenergy 6, 431–439, 2012

Holt et al.Fungal Mycelium and Cotton Plant Materials in the Manufacture of Biodegradable Molded Packaging Material

The largest increase in water absorption from 0.75 h to 3 h

was seen in Liquid 4 which moved from 11.2% to 39.2 %.
The smallest increase from 0.75 h to 3 h was Grain 4 ,
8.9% to 12.2 %.
Table V. Cone calorimeter testing results from molded packaging material produced from six cotton-based substrate blends and two fungal inoculum
methods.
Thermal properties
Response variable Peak heat release rate Average heat release rate Peak carbon monoxide

Standarda ASTM E1354 ASTM E1354 ASTM E1354

Units kW/m2 kW/m2 kW/m2
Inoculum/blend Valueb Inoculum/blend Valueb Inoculum/blend Valueb
Liquid 2 1158A Liquid 2 753A Grain 6 5550 A
Grain 1 1133A Liquid 6 665A B Liquid 2 247 B
Liquid 6 1070A Grain 5 663A B Liquid 4 229 B
Grain 4 1053A 1040A Liquid 4 625A B Liquid 6 213 B

RESEARCHARTICLE
Grain 2 1040A 1035A Grain 3 Liquid 617A B Grain 1 200B C
Grain 3 1033A 991A 5 613A B Grain 5 200B C
Grain 6 973A B Delivered by Ingenta 593A B Liquid 5 197B C
Grain 5 572A B 177B C
919A B toGrain 4 : Liquid 1
D
Liquid 5 565A B Grain 4
Liquid 4 Greg HoltGrain 2 562A B Grain 2 167B C
D
Liquid 1 IP : 553B Grain 3
158B C
199.133.133.66Grain 6 D
Liquid 1 100C D
Thu, 29 Nov 2012
23:20:57Grain 1
Liquid 3 657B Liquid 3 549B Liquid 3 090 D
Response variable Average carbon monoxide Peak carbon dioxide Average carbon dioxide
Standarda ASTM E1354 ASTM E1354 ASTM E1354
Units mg/s mg/s mg/s
Grain 6 5100Ab Grain 6 26180A b Grain 6 24280 A b
Inoculum/blend
Liquid 6 Value
077B Inoculum/blend
Liquid 6 Value
4014B Inoculum/blend
Liquid 2 Value
2465 B
Grain 2 068B C Grain 4 3963B Liquid 6 2203 B
Liquid 4 052B C Grain 1 3897B Liquid 4 2098 B
Liquid 2 050B C Grain 5 3827B C Grain 5 2027 B
Liquid 5 046B C Liquid 2 3826B C Liquid 5 1976 B
Liquid 1 044B C Liquid 5 3412B C Grain 4 1957 B
Liquid 3 034B C Liquid 1 3235B C Grain 1 1863 B
Grain 3 033B C Grain 3 3127B C Liquid 1 1856 B
Grain 1 000C Liquid 4 3119B C Grain 3 1723 B
Grain 4 000C Grain 2 3101B C Liquid 3 1516 B
Grain 5 000C Liquid 3 2378C Grain 2 1446 B
Response variable Peak mass loss rate Average mass loss rate Mass loss
Standard a
ASTM E1354 ASTM E1354 ASTM E1354
Units mg/s mg/s mg/s

Inoculum/blend Valueb Inoculum/blend Valueb Inoculum/blend Valueb

Grain 1 1533A Liquid 3 714A Liquid 4 740 A
Grain 4 1500A B Liquid 6 667A Grain 5 727 A
Liquid 4 1367A B Liquid 5 621A B Grain 1 720 A
Grain 5 1267A B Liquid 4 616A B Grain 2 702 A
Grain 6 1250A B Liquid 1 591A B Grain 6 700 A

J. Biobased Mater. Bioenergy 6, 431–439, 2012 439

 Fungal Mycelium and Cotton Plant Materials in the Manufacture of Biodegradable Molded Packaging Material Holt et al.

Liquid 6 1200A B Grain 6 520A B Grain 4 690 A

Grain 2 1150A B Grain 2 460A B Liquid 2 686 A
Liquid 1 1133A B Grain 1 387A B Liquid 5 555 A
Liquid 5 1067A B Grain 5 370A B Grain 3 527A B
Grain 3 1033A B Grain 3 363A B Liquid 1 462A B
Liquid 3 900A B Grain 4 170A B Liquid 6 396A B
Liquid 2 767B Liquid 2 15B Liquid 3 88 B
Notes: aASTM = American Society for Testing and Materials. bMeans within the same
column followed by different letters in the corresponding row are statistically different at
the 0.05 level of significance.
The percent water absorption after 168 h indicated no
significant differences between any of the treatments. The
largest absorption, after 168 h, was seen in Grain 1 (198%)
with the lowest percent absorption in Grain 3 and Grain 6 ,
93.5% and 94.3%, respectively.

3.3. Thermal Properties

The thermal properties (Tables IV and V) show thermal conductivity 26 and cone calorimetry25 data. The thermal
resistance and conductivity data (Table IV) show Liquid 6 with the highest R-value (1.5) which was signifi-
cantly higher than Grain 1 (0.96), Liquid 4 (0.87), Liquid 5 Fig. 9. Fungal mycelium and cotton plant material molded packaging
material being used by a large office equipment manufacturer in the
(0.84), and Grain 3 (0.82). Thermal conductivity was high-
United States.
est with Grain 3 (0.18) and lowest with Liquid 6 (0.10). The
conductivity values were within the ranges of gypsum (0.17),
high density hardboard (0.15), plywood (0.12), and both
hardwoods (0.16) and softwoods (0.12) at 26.8 C. 32 Cone
calorimeter testing (Table V) showed gas pro-
Delivered by Ingenta to:
duction (carbon monoxide, CO and carbon dioxide, CO2 )
Greg Holt
for Grain 6 to have significantly higher gas production
RESEARCHARTICLE

IP : 199.133.133.66
than all other treatments, 55.5 mg/s—peak CO; 51 mg/s—
average CO; 2618 mg/s peak CO ; 2428 mg/s—average Thu,
2 Cone calorimeter testing showed Liquid 3 had the
CO2. Originally upon examination of the large difference
between the Grain 6 data and the other treatments, the
Grain data was questioned as a potential error in data
entry or perhaps a mistake during the analytical analyses.
Upon further review and analysis, validity of the data was
established by investigation into the interaction of
substrate particle size and method of inoculation to cone
calorimeter analysis. The lowest peak CO gas production
was seen in Liquid 3 (0.9 mg/s). The average CO
production for Grain 1, Grain 4, and Grain 5 show zero
emissions when in actuality they are less than 0.00 mg/s.
With the exception of Grain 6, all other treatments had
average CO production less than 1.0 mg/s. The peak and
average CO2 production was lowest for Liquid 3 (237.8
mg/s—peak; 151.6 mg/s— average) and Grain 2 (310.1
mg/s—peak; 144.6 mg/s— average).
29 Nov 2012 23:20:57
smallest mass loss (8.8%) compared to Liquid 4 with
74%. Liquid 3 had significantly lower mass loss than all
treatments except Grain 3 (52.7%), Liquid 1 (46.2%), and
Liquid 6 (39.5%). The peak and average mass loss rate
were lowest for Liquid 2, 76.7 mg/s and 1.5 mg/s,
respectively. The highest peak and average mass loss rates
were seen in treatments Grain 1 (153.3 mg/s—peak) and
Liquid 3 (71.4 mg/s—average).
3.4. Application of Findings
Results of this study indicate that cotton-based fungal
mycelium packaging material is a viable alternative to
polystyrene packaging. Figures 9 and 10 show two
commercial applications for the type of products produced
from two blends evaluated in this study. Figure 9 shows

440 J. Biobased Mater. Bioenergy 6, 431–439, 2012

Holt et al.Fungal Mycelium and Cotton Plant Materials in the Manufacture of Biodegradable Molded Packaging Material
Fig. 10. Fungal mycelium and cotton plant material molded packaging
material being used by a large computer manufacturer in the United
States.
an application using treatments Grain 1 and Grain 2 and
Figure 10 is a hybrid whose development was a direct
result of the findings in this study.
4. CONCLUSIONS
This study evaluated six cotton-based biomass blends for
use in a process designed to produce an
environmentallyfriendly molded packaging material that
could replace polystyrene packaging currently in the
that presently use fossil-fuel based materials. Delivered by Ingenta to:19. E. Bayer and G. McIntyre, Method for producing
rapidly renewable
Greg Holtchitinous material using fungal fruiting bodies and product made
Abbreviations IP : 199.133.133.66thereby. U.S. Patent Application US2009/0307969 A1, December
Thu, 29 Nov 2012 23:20:
20. (2009)ASTM. C165-07,
marketplace. In addition to the six cotton-based blends,
two methods, grain and liquid, were used to inoculate the
blends with fungal spores resulting in twelve treatments.
The blends were inoculated and the test specimens grown
in tools (plastic molds) for 5 days and then dried to
remove moisture. The recipe for each blend was identical
except for the particle size range of the cotton-based
materials. The difference in the inoculums was that one
carried the fungal spores on kernels of grain whereas the
other had the fungal spores suspended in liquid. The
liquid inoculum was easier to use in the process and
provided a more consistent distribution of fungal spores
when applied to the blends. The grain inoculum generally
resulted in higher densities due primarily to the added
weight of the grain. The densities were higher than
desired (32.04 kg/m3) due in large part to the inclusion of
cotton plant particles less than 2 mm. In future studies,
cotton plant material having a diameter less than 2 mm
will not be used. No single treatment outperformed the
other treatments in all categories evaluated. Most of the
CGB: Cotton Gin Byproducts
Disclaimer
Mention of product or trade names does not constitute an
endorsement by the USDA-ARS over other comparable
J. Biobased Mater. Bioenergy 6, 431–439, 2012 441
treatments performed similarly to each other for the
response variables measured. In regards to percent
degradation associated with accelerated aging testing,
Grain 1 was most consistent in maintaining flexural and
compressive strength and elastic modulus.
Overall, the use of cotton-based fungal mycelium
packaging material is a viable alternative to polystyrene
packaging. As refinements in processing and biomass
blend development continue, the physical and mechanical
properties of the product should improve. Improved phys-
ical characteristics will cause agricultural residue-based
fungal composites to be suitable for numerous
applications
6. S. S. Ahankari, A. K. Mohanty, and M. Misra, Compos. Sci.
57
Standard Test Method for Measuring Compressive
Technol. 71, 653 (2011).
7. H. Gialamas, K. G. Zinoviadou, C.
G. Biliaderis, and
K. P. Koutsoumanis, Food Research International 43, 2402 (2010). 8.
Y. Makino and T. Hirata, Postharvest Biology and Technology 10, 247
(1997).
9. M. Rinaudo, Prog. Polym. Sci. 31, 603 (2006).
10. K. V. H. Prashanth and R. N. Tharanathan, Trends in Food Science
and Technology 18, 117 (2007).
11. I. O. Fasids and U. U. Ekuere, Food Chem. 48, 255 (1993).
12. I. O. Fasids, Food Chem. 55, 161 (1996).
13. A. Philippoussis, R. Diamantopoulou, and C. Israilides, Int.
Biodeterior. Biodegrad. 59, 216 (2007).
14. A. Medina, M. Vassileva, F. Caravaca, A. Roldán, and R. Azcón,
Chemosphere 56, 499 (2004).
15. J. B. Sutherland, Mycologia 76, 369 (1984).
16. Y. Li and K. L. Pickering, Compos. Sci. Technol. 69, 1265 (2009).
17. Y. Li, K. L. Pickering, and R. L. Farrell, Industrial Crops and
RESEARCHARTICLE
Products 29, 420 (2009).
18. E. Bayer, G. McIntyre, and B. L. Swersey, Method for producing
grown materials and products made thereby, U.S. Patent Application
US 2008/0145577 A1, June (2008).
products. Products or trade names are listed for reference
only. USDA is an equal opportunity provider and
employer.
Acknowledgment: This project was a collaborative effort
involving USDA-ARS in Lubbock, TX, and Ecovative
Design, LLC of Green Island, NY, under a
 Fungal Mycelium and Cotton Plant Materials in the Manufacture of Biodegradable Molded Packaging Material Holt et al.

Cooperative Research and Development Agreement 32. F. P. Incropera, D. P. Dewitt, T. L. Bergman, and A. S. Lavine,
(#583K95-0-1391). The authors would like to thank Fundamentals of Heat and Mass Transfer, 6th edn., John Wiley &
Philip Sons, Inc., Hoboken (2007).

Pearson, Bill Turner, Jeff Turner, Jimmie Castro, Gary

Schlabs, Clinton Cox, and Chris Arinder for their work in Received: 28 July 2011. Accepted: 2 April 2012.
preparation of this manuscript, processing the cotton
byproducts, and making the blends used in this study.

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