ALTERNATIVE MUSHROOM PRODUCTION SYSTEMS USING NON-COMPOSTED GRAIN ...

253 Pages·2007·2.04 MB·English

The Pennsylvania State University
The Graduate School
Department of Agricultural and Biological Engineering

ALTERNATIVE MUSHROOM PRODUCTION SYSTEMS USING NON-
COMPOSTED GRAIN-BASED SUBSTRATES

A Thesis in
Agricultural and Biological Engineering
by
Mark A. Bechara

©2007 Mark A. Bechara
Submitted in Partial Fulfillment
of the Requirements
for the Degree of
Doctor of Philosophy
December 2007
The thesis of Mark A. Bechara was reviewed and approved* by the following:

Paul H. Heinemann
Professor of Agricultural and Biological Engineering
Thesis Advisor
Chair of Committee

Paul N. Walker
Professor of Agricultural and Biological Engineering

Tom L Richard
Associate Professor of Agricultural and Biological Engineering

John M Regan
Assistant Professor of Environmental Engineering

C. Peter Romaine
Professor of Plant Pathology

Roy E. Young
Professor of Agricultural and Biological Engineering

Head of the department of Agricultural and Biological Engineering

*Signatures are on file at the Graduate School

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ABSTRACT

The commercial production system of Agaricus bisporus mushroom relies
entirely on composting as a means to generate a mushroom-specific substrate, and all
aspects of this traditional system are designed for the preparation, processing, and
handling of this compost-based substrate. Furthermore, the substrate, which is a mixture
of compost and delayed-release supplements, has been refined for maximum mushroom
production, and although widely adopted by mushroom producers around the world, this
system is environmentally problematic (odor emissions, nutrient-rich run-off, and
substrate disposal limitations) and these problems will only intensify with urban sprawl.
In this study, grain-based mushroom production systems were evaluated as a
potential alternative to the traditional compost-based system for A. bisporus. Two types
of grain-based systems were developed and called– “Satellite Mushroom Production
System” (SMPS), and “Complete On-site Mushroom Production System” (COMPS).
Mushroom producers adopting the SMPS, would use a substrate primarily composed of
commercial grain spawn, whereas mushrooms producers adopting the COMPS would use
a subsrate composed of cereal grains mixed with oilseed (grain/oilseed). Both systems
were tested and refined for mushroom production and substrate bioefficiency (BE).
Furthermore, the theoretical designs and the direct costs for mushroom production for
each system were developed and calculated.
For the SMPS, the factors influencing mushroom production were: adding a layer
of water-holding materials below the commercial grain spawn substrate, heat-sterilizing
casing with the addition of activated carbon, and type and level of delayed-release
supplement used. Overall, the highest yield of mushrooms for a commercial grain spawn
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substrate was 14.28 kg/m2 with a corresponding BE of 177% and was obtained for a
treatment containing 5% S41- an underlying layer of perlite (2000 ml) and cased with a
heat-sterilized casing containing 25% activated carbon. The design of SMPS entails
different steps that start with mixing of the substrate ingredients, transferring the
substrate to trays, and subsequent steps are comparable to traditional compost-based
systems. Based on the cost model developed for the SMPS, the cost of production/unit
weigh of mushroom is $4.00/kg. This is much higher than the market cost of A. bisporus
and mushroom producers would not make a profit.
For the COMPS, the factors that were tested for mushroom production using
grain/oilseed substrate were type of cereal grain used, type and rate of oilseed added, pre-
conditioning the substrate with Scytalidium thermophilum (a mushroom compost
thermophile), and moisture content in the grain portion of the substrate. Furthermore, two
additional mushroom producing species (A. blazei and Pleurotus eryngii) were grown on
the grain/oilseed substrate to determine whether the grain-based system could be
extended to the production of other mushroom forming species.
For A. bisporus, adding oilseeds to a basal substrate composed of millet grain
increased yield and a 15% amendment of soybean gave the highest yield (16.9 kg/m2
with a corresponding BE of 205%). Furthermore, preconditioning the substrate with S.
thermophilum improved yield on oat based-substrates and decreased spawn-run time for
all grain/oilseed substrates. Moisture contents ranging from 50-65% seem adequate for
mushroom production, whereas the peak in yield is observed between the 55% and 60%
moisture levels.
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For A. blazei and P. eryngii mushroom production, grain/oilseed substrate
successfully supported mushroom production. For A. blazei, millet with 30% niger
yielded 15.9 kg/m2, whereas, an oat/oilseed substrate provided the highest yield of 106 g
for P. eryngii. Overall, the highest yield for A. bisporus was observed for a millet/ 5%
soybean substrate with an additional amendment of 5% delayed-release supplement
(Promycel Target®) which produced 21.3 kg/m2 with a BE of 273%. The system design
is composed of an aseptic processing unit that converts the raw materials into a suitable
mushroom substrate, and steps following the processing of the grain/oilseed substrate are
comparable to the SMPS. Based on the cost model developed for the COMPS, the cost of
production/unit weight of mushrooms is $1.40/kg, and this would provide mushroom
producers with a profit.
Based on findings from this research, developing an alternative commercial A.
bisporus mushroom production system using grain-based substrates is a highly promising
alternative to commercial compost-based system and its environmental problems. Yield
from grain-based substrates is still lower than compost-based substrates but further
refinement of the substrate ultimately will increase mushroom yield. A pilot-scale system
is needed to test scale-up of this system for mushroom production on grain-based
substrates.

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TABLE OF CONTENTS

LIST OF FIGURES......................................................................................................................................X
LIST OF TABLES....................................................................................................................................XII
ACKNOWLEDGMENTS.......................................................................................................................XIV
1. INTRODUCTION....................................................................................................................................1
2. LITERATURE REVIEW........................................................................................................................5
2.1. AGARICUS BISPORUS MUSHROOM FUNGUS.........................................................................................5
2.2. SPAWN PRODUCTION...........................................................................................................................6
2.3. NUTRIENT SUPPLEMENTS USED BY MUSHROOM PRODUCERS.............................................................8
2.4. COMMERCIAL PRODUCTION OF AGARICUS BISPORUS..........................................................................9
2.4.1. Composting in Commercial Agaricus bisporus Production........................................................9
2.4.2. Mushroom Fungus Spawning....................................................................................................12
2.4.3 Substrate Casing and Casing Inoculum.....................................................................................13
2.4.4. Mushroom Fungus Pinning and Harvesting.............................................................................15
2.4.5. Metabolism and Physiology of A. bisporus...............................................................................15
2.5. AGARICUS MUSHROOM PRODUCTION ON NON-COMPOSTED SUBSTRATES........................................19
2.6. AGARICUS BISPORUS USED IN ANIMAL FEED....................................................................................21
2.7. SOLID STATE FERMENTATION...........................................................................................................22
2.7.1. Definition of Solid State Fermentation......................................................................................22
2.7.2. Support Material Used in SSF..................................................................................................23
2.7.3. Grains and Oilseeds..................................................................................................................24
2.7.4. Environmental and Process Parameters in SSF.......................................................................26
2.7.5. Fungal Biomass Estimation in SSF...........................................................................................28
2.8. ASEPTIC PROCESSING OF PARTICULATE MATERIALS.........................................................................30
2.8.1. Gelatinization of Starch............................................................................................................30
2.8.2. Thermal Processing and its Application to Food......................................................................31
2.8.3. Steam-based Thermal Sterilization Processes..........................................................................31
2.8.4. Aseptic Processing History and Description............................................................................32
2.8.5. Elements of a Continuous Aseptic Processing System..............................................................34
2.8.5.1. Heating Section......................................................................................................................34
2.8.5.2. Holding Section......................................................................................................................34
2.8.5.3. Cooling Section......................................................................................................................34
2.8.5.4. Aseptic Filling and Packaging Section..................................................................................35
2.8.6. Liquid-based Segmented Flow Aseptic Processing System.......................................................35
2.8.7. Steam-based Segmented-flow Processing System.....................................................................36
2.9. CONTINUOUS PRODUCTION OF MUSHROOM SPAWN..........................................................................37
2.10. SUMMARY OF LITERATURE REVIEW................................................................................................38
CHAPTER 3 PRODUCTION OF AGARICUS BISPORUS MUSHROOMS ON COMMERCIAL GRAIN
SPAWN MIXED WITH S41 AND S44 SUPPLEMENTS..........................................................................39
3.1 ABSTRACT..........................................................................................................................................39
3.2. INTRODUCTION..................................................................................................................................40
3.3. METHODS..........................................................................................................................................44
3.3.1. Mushroom Grain Spawn, Nutrient Supplement and Water-holding Material..........................44
3.3.2. Preparation of Mushroom Production Containers...................................................................44
3.3.3. Mushroom Production Chamber...............................................................................................45
3.3.4. Summary of the Experimental set-up........................................................................................45
3.3.5. Parameters Evaluated and Analysis of Data............................................................................46
3.4. RESULTS............................................................................................................................................46
3.5. DISCUSSION.......................................................................................................................................49
3.6. CONCLUSIONS....................................................................................................................................52
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CHAPTER 4 FACTORS INFLUENCING MUSHROOM YIELD IN NON-COMPOSTED
COMMERCIAL GRAIN SPAWN SUBSTRATES.....................................................................................54
4.1. ABSTRACT.........................................................................................................................................54
4.2. INTRODUCTION..................................................................................................................................55
4.3. METHODS..........................................................................................................................................58
4.3.1. Mushroom Hybrid Nutrient Supplement and Water-holding Material.....................................58
4.3.2. Preparation of Mushroom Production Containers...................................................................58
4.3.3. Environmental Conditions in Mushroom Production Rooms...................................................59
4.3.4. Experimental Set-up and Analysis of Data...............................................................................60
4.4. RESULTS............................................................................................................................................61
4.5. DISCUSSION.......................................................................................................................................67
4.6. CONCLUSIONS....................................................................................................................................68
CHAPTER 5 EFFECT OF DELAYED-RELEASE SUPPLEMENTS IN GRAIN-BASED SUBSTRATE
ON YIELD OF THE MUSHROOM (AGARICUS BISPORUS)..................................................................70
5.1. ABSTRACT.........................................................................................................................................70
5.2. INTRODUCTION..................................................................................................................................71
5.3. METHODS..........................................................................................................................................74
5.3.1. Mushroom Grain Spawn Supplements and Water-holding Material........................................74
5.3.2. Preparation of Mushroom Production Containers...................................................................74
5.3.3. Settings in Mushroom Tray Reactor.........................................................................................75
5.3.4. Summary of the Experimental Design.......................................................................................75
5.3.5. Parameters Evaluated and Analysis of Data............................................................................75
5.4. RESULTS............................................................................................................................................76
5.4.1 Mushroom Yield.........................................................................................................................76
5.4.2. Substrate Bioefficiency..............................................................................................................79
5.4.3. Mean Mushroom Weight...........................................................................................................79
5.4.4. Substrate Temperature..............................................................................................................79
5.5. DISCUSSION.......................................................................................................................................81
5.6. CONCLUSIONS....................................................................................................................................83
CHAPTER 6 EVALUATING THE ADDITION OF ACTIVATED CARBON TO HEAT TREATED
MUSHROOM CASING FOR GRAIN-BASED AND COMPOST-BASED SUBSTRATES.....................85
6.1. ABSTRACT.........................................................................................................................................85
6.2. INTRODUCTION..................................................................................................................................86
6.3. METHODS..........................................................................................................................................89
6.3.1. Spawn, Nutrient Supplement and Substrate..............................................................................89
6.3.2. Casing Preparation...................................................................................................................89
6.3.3. Preparation of Mushroom Trays...............................................................................................90
6.3.4. Conditions in Tray Bioreactor..................................................................................................90
6.3.5. CO Absorption Test.................................................................................................................91
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6.3.6. Experimental Design.................................................................................................................92
6.4. RESULTS............................................................................................................................................93
6.5. DISCUSSION.......................................................................................................................................98
6.6. CONCLUSIONS..................................................................................................................................102
CHAPTER 7 CULTIVATION OF AGARICUS BISPORUS AND AGARICUS BLAZEI ON
SUBSTRATES COMPOSED OF CEREAL GRAINS AND OILSEEDS.................................................104
7.1 ABSTRACT........................................................................................................................................104
7.2. INTRODUCTION................................................................................................................................105
7.3. MATERIALS AND METHODS.............................................................................................................109
7.3.1. Fungal Strains, Seeds, and Supplements.................................................................................109
7.3.2. Grain Substrate Preparation Process.....................................................................................109
7.3.3. Mushroom Production Tray Preparation...............................................................................110
7.3.4. Tray Bioreactor Conditions....................................................................................................111
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7.3.5. Measured Variables in Tray Bioreactors................................................................................111
7.3.6. Experimental Set-up and Statistical Analysis..........................................................................112
7.4. RESULTS..........................................................................................................................................113
7.4.1. Production Time......................................................................................................................113
7.4.2. Environmental Conditions in Tray Bioreactors......................................................................113
7.4.3. Agaricus bisporus...................................................................................................................115
7.4.4. Agaricus blazei........................................................................................................................119
7.5. DISCUSSION.....................................................................................................................................121
7.6. CONCLUSIONS..................................................................................................................................124
CHAPTER 8 PRE-INCUBATING NON-COMPOSTED GRAIN SUBSTRATES WITH THE
THERMOPHILIC FUNGUS SCYTALIDIUM THERMOPHILUM FOR MUSHROOM (AGARICUS
BISPORUS) PRODUCTION......................................................................................................................126
8.1. ABSTRACT.......................................................................................................................................126
8.2. INTRODUCTION................................................................................................................................127
8.3. METHODS AND MATERIALS.............................................................................................................130
8.3.1. Fungal Cultures Substrate Materials......................................................................................130
8.3.2. Preparation of Grain Substrates.............................................................................................131
8.3.3. Preparation of Mushroom Production Containers.................................................................131
8.3.4. Environmental Conditions in Mushroom Production Chamber.............................................132
8.3.5. Experimental Set-up and Analysis of Data.............................................................................133
8.3.6. Oxygen Uptake Rate Evaluation.............................................................................................135
8.4. RESULTS..........................................................................................................................................136
8.4.1. Preliminary Incubation experiments with S. thermophilum....................................................136
8.4.2. Treatments with Shorter Incubation Durations with S. thermophilum...................................137
8.4.3. Oxygen Uptake Rate Evaluation.............................................................................................139
8.5. DISCUSSION.....................................................................................................................................140
8.6. CONCLUSIONS..................................................................................................................................142
CHAPTER 9 EFFECT OF SUBSTRATE MOISTURE CONTENT ON AGARICUS BISPORUS
OXYGEN CONSUMPTION RATE, HYPHAL EXTENSION RATE, MUSHROOM YIELD, AND
BIOEFFICIENCY......................................................................................................................................144
9.1. ABSTRACT.......................................................................................................................................144
9.2. INTRODUCTION................................................................................................................................145
9.3. METHODS........................................................................................................................................148
9.3.1. Fungal Cultures and Substrate Materials...............................................................................148
9.3.2. Substrate Preparation for OUR Measurements and Mushroom Fruiting...............................149
9.3.3. Procedure for OUR Measurement..........................................................................................150
9.3.4. Oxygen Uptake Rate (OUR) Calculation................................................................................151
9.3.5. Dry Matter and Ash Measurements........................................................................................152
9.3.6. Linear Extension Rate Measurements.....................................................................................152
9.3.7. Experimental Set-up................................................................................................................153
9.4. RESULTS..........................................................................................................................................154
9.4.1. OUR Measurements as affected by the Mass of Substrate and NaOH Pellets........................154
9.4.2. OUR Readings as Affected by Substrate Moisture Content....................................................155
9.4.3. OUR Readings as Affected by Measuring Frequency.............................................................160
9.4.4. Mycelium Extension Rate........................................................................................................161
9.4.5. Yield and Substrate Bioefficiency as Affected by Substrate Moisture Content.......................163
9.5. DISCUSSION.....................................................................................................................................164
9.6. CONCLUSIONS..................................................................................................................................167
CHAPTER 10 TWO ALTERNATIVE AGARICUS BISPORUS MUSHROOM PRODUCTION
SYSTEMS USING GRAIN-BASED SUBSTRATES...............................................................................168
10.1 ABSTRACT......................................................................................................................................168
10.2. INTRODUCTION..............................................................................................................................169
10.2.1. Steps in Agaricus bisporus Mushroom Production...............................................................172
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10.2.2. Factors Influencing Mushroom Yield in Grain-based Substrates.........................................175
10.3. DESCRIPTION OF ALTERNATIVE MUSHROOM PRODUCTION SYSTEM.............................................176
10.3.1 Satellite Mushroom Production System.................................................................................176
10.3.2. Complete On-site Mushroom Production System.................................................................177
10.3.3. Duration of Mushroom Production Process.........................................................................182
10.4. COST MODEL FOR GRAIN-BASED MUSHROOM PRODUCTION SYSTEM...........................................183
10.4.1. Cost Model for Satellite Mushroom Production System.......................................................184
10.4.2. Cost Model for Complete On-site Mushroom Production System........................................188
10.5. DISCUSSION...................................................................................................................................192
10.6. CONCLUSIONS................................................................................................................................194
CHAPTER 11 CONCLUSIONS AND SCOPE FOR FUTURE RESEARCH.........................................195
REFERENCES.........................................................................................................................................200
APPENDIX A CULTIVATION OF PLEUROTUS ERYNGII ON SUBSTRATES COMPOSED OF
GRAINS AND OILSEEDS........................................................................................................................220
APPENDIX B RYE GRAIN SUBSTRATE FLOWABILTY...................................................................226
APPENDIX C COST MODEL EXCEL SPREADSHEETS.....................................................................232
VITA................................................................................................................................................................

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List of Figures

Figure 2.1. Segmented-flow aseptic processing schematic (Walker and Beelman, 2002).
...................................................................................................................................36
Figure 2.2. Continuous steam segmented-flow, aseptic processing system (Anderson and
Walker, 2005)...........................................................................................................37
Figure 3.1. Effect of adding a single layer of perlite below the commercial grain spawn
substrate containing 10% S41 on mushroom production.........................................49
Figure 4.1. Summary of the main effects (S41 rate, perlite volume, and casing type) on
mushroom yield (kg/m2)...........................................................................................63
Figure 4.2. Summary of the main effects (S41 rate, perlite volume, and casing type) on
BE (%)......................................................................................................................64
Figure 4.3. Summary of significant interaction effects of S41 rate, perlite volume, and
casing type on mushroom yield (kg/m2)...................................................................66
Figure 5.1. Mushroom yields for non-composted substrates composed of commercial
millet grain spawn and different rates of five delayed-release supplements............77
Figure 5.2. Comparison of the temperature profiles during the 23-day period after casing
for the production room (Env.) and non-composted substrates composed of
commercial millet grain and five different five delayed-release supplements.........80
Figure 6.1. Change in pH of peat moss heat-treated for different time intervals in an
autoclave...................................................................................................................93
Figure 6.2 pH for different casing treatments, before and after autoclaving....................94
Figure 6.3. Comparison between activated carbon and NaOH as carbon dioxide
absorbent.................................................................................................................100
Figure 7.1. Environmental conditions and mushroom yield/day for A. bisporus tray
bioreactor................................................................................................................114
Figure 7.2. Environmental conditions and mushroom yield/day for A. blazei tray
bioreactor................................................................................................................115
Figure 7.3.Yield from two millet-grain substrates prepared on-site with one treatment
receiving stage II supplementation, and compared to commercial grain spawn
(Sylvan) substrate with stage II supplementation...................................................116
Figure 7.4. A. bisporus fruiting on a substrate composed of grain and oilseed..............118
Figure 7.5. A. blazei mushroom fruiting on a grain/oilseed substrate............................121
Figure 8.1. OUR measurements for A. bisporus and S. thermophilum at 16, 24, and 32oC.
.................................................................................................................................140
Figure 9.1. OUR observations as affected by the mass of the substrate with the red-line
indicating OUR when all the observations are average..........................................155
Figure 9.2. OUR of A. bisporus growing on a rye grain substrate with different substrate
moisture contents treatments with rye grain spawn................................................157
Figure 9.3. Different moisture level treatments for a rye grain-based substrate inside the
Oxtop® bottles........................................................................................................158
Figure 9.4. Change in porosity for the different rye grain substrate at different moisture
levels at the beginning and at the end of the OUR measurement that spanned 32
days.........................................................................................................................159
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