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Cellulose-Mycelia foam : novel bio-composite material Ahmadi, Hoda 2016

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Cellulose-Mycelia Foam: NovelBio-Composite MaterialbyHoda AhmadiB.Sc. Chemical Engineering, Amirkabir University of Technology, 2012A THESIS SUBMITTED IN PARTIAL FULFILLMENT OFTHE REQUIREMENTS FOR THE DEGREE OFMASTER OF APPLIED SCIENCEinThe Faculty of Graduate and Postdoctoral Studies(Mechanical Engineering)THE UNIVERSITY OF BRITISH COLUMBIA(Vancouver)December 2016c© Hoda Ahmadi 2016AbstractDemand for sustainable products is growing faster than ever before. Because of this,the development of novel sustainable materials is crucial to leverage our environmentalresources and to ensure future growth of Canada’s economy. In this study, we propose atechnology to develop the use of fungal mycelium, the vegetative part of a fungus, througha porous scaffold of cellulose-based foam. A methodology for producing cellulose-myceliafoam (CMF) has been developed by mixing a surfactant with pulp suspension of 1%consistency and Pleurotus djamor spawn, mixing at high velocity to entrain air, filteringthe suspension, and then holding at incubation conditions suitable for mycelium growth.During the incubation period, temperature (20-25 ◦C), pH (5-8), humidity (80-100%),ventilation and exposure to light were controlled. Simplicity of production, biodegrad-ability, and 3-D porous structure of the product position this biocomposite as a greenalternative to polymeric foams. The structure of the CMF was characterized throughfluorescent microscopy during the incubation period. The effect of mycelial growth onthe mechanical behaviour of the CMF including compressibility, thermal decomposition,dry and wet strength was investigated during 25 days of mycelial growth. The resultsindicated that all tested mechanical properties improved after 25 days of mycelial growth.The second set of experiments was run to specify the application of the CMF in a hy-draulic filtration system. The pressure drop, permeability, and filtration efficiency ofthe product were studied. The experimental results showed that the permeability of theCMF decreases by an increase in mycelial growth. The hydraulic filtration efficiency ofthe product improved from 74% for cellulosic foam to 99.9% for 25 days CMF for remov-ing 20 µm and larger particles. Bioremediation tests also were performed to evaluate thedetoxification capability of mycelia in the CMF. Detoxification tests demonstrated thatthe living mycelia are able to detoxify potassium hydroxide from waste alkaline batteries.iiPrefaceThe work in this dissertation, including all experiments, analysis and writing was con-ducted by the author, H. Ahmadi. All work was performed under supervision of Dr.James A. Olson.iiiTable of ContentsAbstract . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . iiPreface . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . iiiTable of Contents . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . ivList of Tables . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . viList of Figures . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . viiAcknowledgements . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . ixDedication . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . x1 Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11.1 Motivation and advantages of biodegradable composites . . . . . . . . . 11.2 Background . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 21.2.1 Foam-paper . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 21.2.2 Mycelium and sustainability . . . . . . . . . . . . . . . . . . . . 71.2.3 Filter media . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 111.3 Cellulose-Mycelia Foam as a filtration media . . . . . . . . . . . . . . . 132 Cellulose-Mycelia Foam: production and characteristics . . . . . . . . 142.1 Production . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 142.2 Morphology and thermo-mechanical properties . . . . . . . . . . . . . . 172.2.1 Morphology . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 17ivTable of Contents2.2.2 Thermal Gravimetric Analysis . . . . . . . . . . . . . . . . . . . 192.2.3 Dynamic Mechanical Analysis . . . . . . . . . . . . . . . . . . . . 212.2.4 Tensile and wet strength . . . . . . . . . . . . . . . . . . . . . . . 233 Applications . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 273.1 Water filtration . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 273.1.1 Background . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 273.1.2 Methods . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 283.1.3 Results and discussion . . . . . . . . . . . . . . . . . . . . . . . . 293.2 Mycoremediation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 343.2.1 Background . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 343.2.2 Methods . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 363.2.3 Results and discussion . . . . . . . . . . . . . . . . . . . . . . . . 364 Conclusions and future work . . . . . . . . . . . . . . . . . . . . . . . . . 39Bibliography . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 41vList of Tables1.1 Role of mycelium in degradation of pollutants. . . . . . . . . . . . . . . . 91.2 Role of mycelium in the removal of pollutants by using biosorption process. 102.1 Characteristics of the cellulose-mycelia foam samples. . . . . . . . . . . . 153.1 Selected Beta ratio and the corresponding efficiencies of the CMF filtersfor 5 µm, 10 µm, and 20 µm. . . . . . . . . . . . . . . . . . . . . . . . . 323.2 An average composition of Alkaline battery . . . . . . . . . . . . . . . . . 35viList of Figures1.1 NBSK foam-paper at 50% air content. . . . . . . . . . . . . . . . . . . . 31.2 Image of mycelial strands . . . . . . . . . . . . . . . . . . . . . . . . . . . 82.1 Image of dried cellulose-mycelia foam after 25 days. . . . . . . . . . . . . 142.2 Fluorescent micrographs and conventional photographs showing the effectof mycelial growth on the microstructure of cellulose-mycelia foam. . . . 182.3 The effect of mycelial growth on the thermal stability of the cellulose-mycelia foam. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 192.4 TGA curve and first derivative of the weight loss for different stages ofmycelial growth. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 202.5 The effect of mycelial growth on the compressibility of the cellulose-myceliafoam. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 222.6 The effect of mycelial growth on the tensile index and wet strength of thecellulose-mycelia foam. . . . . . . . . . . . . . . . . . . . . . . . . . . . . 242.7 Interaction between fibres in foam-paper and cellulose-mycelia foam. . . . 263.1 The hydraulic circuit for measuring the pressure drop of the CMF filters. 283.2 The effect of mycelial growth on the pressure drop of the cellulose-myceliafoam under continuous loading of water. . . . . . . . . . . . . . . . . . . 303.3 Schematic view of a 1-D model of the filtration apparatus used to calculatethe permeability of the CMF samples. . . . . . . . . . . . . . . . . . . . . 303.4 The effect of mycelial growth on the normalized hydraulic permeability ofthe cellulose-mycelia foam. . . . . . . . . . . . . . . . . . . . . . . . . . . 313.5 The effect of mycelial growth on the hydraulic filtration efficiency of thecellulose-mycelia foam. . . . . . . . . . . . . . . . . . . . . . . . . . . . . 32viiList of Figures3.6 Quality factor of the cellulose-mycelia foam as a hydraulic filter media. . 333.7 Market share for batteries, based on tonnes sold in Canada . . . . . . . . 353.8 Detoxification of KOH by dead and live mycelia of P. djamor in contami-nated water during 60 days. . . . . . . . . . . . . . . . . . . . . . . . . . 373.9 Biosorption mechanism according to the dependence on the cell metabolism. 38viiiAcknowledgementsI would like to express my deepest appreciation to my supervisor, Professor James A.Olson, and my co-supervisor, Professor D. Mark Martinez who continually and convinc-ingly conveyed a sprit of adventure in regard to research. Without their guidance andpersistent help this dissertation would have not been possible.I would specially like to thank Pouyan Jahangiri, Amir Farzad Forughi, Amin En-garnevis, David Sommer, Amirhossein Salimian, and Mohammad Shanbghazani for theirinvaluable discussions and for their patience with my endless stream of questions.I would like to acknowledge the invaluable assistance of Dr. Reza Korehei, GeorgeSoong, Anna Jamroz, Meaghan Miller, and Emilia Jahangir at the UBC Pulp and PaperCentre for assisting me with performing the experiments.I would also like to thank the Natural Science and Engineering Research Council ofCanada (NSERC) and all members of Green Fibre Networks society for their financialsupport.Most of all, I would like to express my heartfelt gratitude to my wonderful parentsMahin and Rostam whom this dissertation is dedicated to, and my lovely brother, Avesta.I have always felt them beside me in Canada, even though they live far away.ixDedicationTo my beloved parentsfor their unconditional love, support, and encouragement.xChapter 1Introduction1.1 Motivation and advantages of biodegradablecompositesComposite materials are engineered materials made from two or more constituents withsignificantly different mechanical properties. There are two categories of constituentmaterials: matrix and reinforcement. The matrix surrounds and supports the reinforce-ments by maintaining their relative positions, while the reinforcement material providesthe desired physical properties. An enormous breadth of materials are used for matricesand reinforcements, depending on the application [1].The rapid growth in the economy, the extraction of natural resources for manufac-turing and production, and the patterns of consumers consumption are the main causeof environmental deterioration [2]. Concern over the environment has evolved from the1960s ecology movement. As the environment continues to worsen, it has become apersistent public concern in developed countries and has recently awakened developingcountries to the green movement. Also, the high rate of depletion of petroleum resourceshas decreased pressures for the dependence on petroleum products while increasing in-terest in maximizing the use of renewable alternatives. Another primary limitation ofusing synthetic polymer composites is that their recycling is quite difficult, and it is oftenpreferred to produce new materials rather than recycling. Consequently, time resistantpolymeric wastes are becoming highly unacceptable, necessitating the search for alter-native materials to existing polymers. The concept of green composites from naturalresources emerged to help society achieve sustainable consumption [3]. The word greenrefers to those materials which are biodegradable, renewable, and sustainable with lowerenvironmental impact. Green composites can be disposed of or composted without harm-11.2. Backgrounding the environment [4]. Polymers from natural resources such as cellulose, starch, lignin,cellulose acetate, poly-lactic acid (PLA), polyhydroxylalkanoates (PHA) are degradableand are classified as biopolymers [5]. The development of biocomposites from sustain-ably sourced biodegradable polymers and natural fibres have attracted great interest inthe composite material sciences as a viable alternative to petroleum based composites[6–10]. While many biocomposites boast a natural reinforcement, the matrix is oftenbased on petrochemicals. A renewable alternative for these matrices is the vegetativestate of a fungus, the mycelium [11]. In this study, the cellulose fibres were used as anatural reinforcement, and the mycelium acted as a bio-based matrix to develop a novelgreen biocomposite.1.2 BackgroundTo better understand the concept of cellulose-mycelia foam, it seems appropriate to havea brief review of cellulose foam and mycelium. A brief background of foam formingtechnology and its application in the papermaking industry is presented in this section,as well as an introduction to mycelium and its application in filtration and bioremedia-tion. This section also gives a brief review of nonwoven hydraulic filter media to help tounderstand filtration mechanisms and the applications of fibrous filters.1.2.1 Foam-paperUltralight 3D porous materials have many important applications in the fields of soundand energy absorption, thermal insulation, radiation shielding, and filtration [12–17].Most of these ultralight materials such as metal foams and aerogels, have traditionallybeen developed using either expensive materials or complicated procedures which haslimited their commercial feasibility and widespread adoption. Another class of materialswhich are used in foam forming technology are cellulose containing materials. Jahangiri etal. [18, 19] recently developed a lightweight, highly porous and 3D shaped cellulose-basedmaterial, referred to as a foam-paper, as shown in Figure 1.1. The production of foam-paper shares many similarities with the papermaking process. Foam-paper offers several21.2. Backgroundkey advantages over papermaking technology, including prevention of fibre flocculation,a 3D porous structure and less water and energy consumed in the manufacturing process.Foam-paper can be used in a wide range of applications including insulation, packaging,filtration and acoustics [18, 20]. A brief review of the foam forming process and itsapplication in the papermaking industry is presented in the next section.Figure 1.1: NBSK foam-paper at 50% air content. (Photographed by Anna Jamroz,UBC Pulp and Paper Centre)Foam forming: state of the artThe idea of applying foam in the papermaking process was first proposed by Radvan andGatward [21] in 1972 and Smith and Punton [22] in 1974 to improve the uniformity of thepaper. Radvan and Gatward [21] used foam as a means of preventing fibre flocculationin the papermaking process due to the presence of very long fibres or high consistencyof pulp suspension. They also developed a foam forming process, referred to as theRadfoam process which employs a discontinuous foam forming unit attached to a smallpaper machine. Also, another study on the Radfoam process [23] showed that the specificvolume (bulk) of a Radfoam-made sheet is 20 to 30 percent higher than a standardhandsheet. Smith et al. [22] and Tringham [23] also confirmed that paper has a higheruniformity, porosity and strength by using the Radfoam process. Another study by Smithet al. [24] on the structure and characteristics of the Radfoam-made sheets showed that31.2. Backgroundsurface tension and bubble spacing in the foam affect the properties of the final product,while chemical effects were found to be insignificant. Recently, Al-Qararah et al. [25]at VTT Technical Research Centre of Finland studied the effect of various parameterson bubble size distribution in foam-formed cellulose fibres. They used a CCD cameraand image processing to characterize the bubble size distribution as a function of variousparameters, such as air content, type and concentration of surfactant, and rotationalspeed. They found that the average bubble size decreases by increasing the rotationalspeed of mixing; also, by decreasing air content, the average bubble radius increases. Theresults showed that the pore size distribution of paper made with foam forming processdepends on the properties of the foam.Recently, with an increasing demand for green and environmentally friendly products,interest in re-applying foaming process for developing novel cellulose-based material hasincreased [18, 26–29]. Many studies have been done on forming lightweight porous mate-rial from nanofibrillated cellulose (NFC) by applying several different methods [26–29].Deng et al. [27] developed a procedure to produce nanoporous cellulose foams by dis-solving cellulose in ionic liquid and then freeze drying the solution. The results showedthat the foam has a 3D open fibrillar network structure with high porosity and highspecific surface area. They also found that cellulose concentration and drying techniquesaffect the structure and pore size of the foam. Sehaqui et al. [28] and Mohammad Aliet al. [29] studied characteristics, morphological structure and mechanical properties ofNFC foams. They found that the foams have a unique cellular structure and the mechan-ical properties of the foams can be improve by controlling the cell structure. In 2012,Cervin et al. [26] proposed a new foam forming process by using the Pickering emulsiontechnique to prepare low density cellulose material. NFC was used to stabilize the airbubbles in an aqueous suspension. In this process, NFC stabilized foam was preparedby adding octylamine to NFC, then the mixture was rapidly agitated for 20 min. Theresulting mixture was foamed with a stainless steel milk beater for 10 min. The NFCstabilized foam was then filtered by Buchner funnel to drain any excess water. Finally,the foam was dried at ambient conditions. The results showed that the porosity of NFCstabilized foam is higher than other cellulose foams made by freeze-drying, while its me-chanical properties are the same as cellulose aerogels and other cellulose foams made41.2. Backgroundby freeze-drying. Recently, Madani [19] and Jahangiri [18] developed a manufacturingprocess which has many similarities with the Radfoam process to produce a lightweightporous cellulose-base foam, referred to as a foam-paper. In this process, surfactant isadded to the pulp/water suspension at pH of 7. Microbubbles are created by agitatingthe mixture and the suspension is filtered by a Buchner funnel to drain any excess water.Two techniques of drying were applied on the wet foam: air drying without pressingand freeze drying. They studied the morphology, mechanical behaviour and prospectiveapplications of the foam-paper for different pulp types and different drying methods.Foam-paper characteristicsWhen first developed, foam-paper was a novel cellulose based product which requiredmore specific studies to characterize its properties. Madani [19] and Jahangiri [18] puta great effort into studying the properties and behaviour of foam-paper. They foundthat density of the foam depends on two parameters: air content and cellulose fibrelength. The results showed that by increasing fibre length, the values of bulk increaseswhich leads to lower density and lower tensile index (TI). They also found that foamair content has a similar effect on bulk, density and TI. To improve the strength ofthe final product, various ratio of refined fibres were added to the suspension. Theresults showed that regardless of the ratio of refined fibres in the foam-paper samples, allsamples exhibited improved mechanical properties as compared to hardwood samples.They found that longer fibres contribute to achieving low density while fine or highlyfibrillated fibres contribute to the strength by diminishing the values of bulk. The effectof pulp consistency on the properties of foam-paper has been studied [30]. The resultsshowed that the density of the foam increases by increasing the pulp consistency whichleads to the lower values of bulk. The effect of air content on the porosity 1 of theproduct has been studied [18, 19]. The results showed that by increasing air contentof the foam, the porosity of final product increases up to 99.7%. Another significantmechanical property is permeability which is the ability of the foam to allow fluid to passthrough it. The results showed that increased bulk, indicative of a more open structure1The ratio of volume of pores to the total volume of sample.51.2. Backgroundwhich results in higher permeability to air flow [19].Korehei et al. [30] studied three different drying techniques to compare the effect ofdrying method on foam-paper characteristics. Drying methods which were applied on thefoam-paper were vacuum dewatering followed by air drying (VAD), vacuum dewateringfollowed by freeze drying (VFD), and direct freeze drying (FD). The morphological studyby SEM and stress-strain curve of the foam-paper by Dynamic Mechanical Analysis(DMA) revealed that the microstructure of foam-paper was affected by applying differentdrying techniques. The VAD samples had the most compact structure due to shrinkingduring the vacuuming and dewatering process; while the FD samples had the highestporosity and the lowest density compare to VAD and VFD samples.Foam-paper applicationsThe 3D structure, high porosity, and low pressure drop of foam-papers make these mate-rial appropriate for a wide range of applications including filtration, packaging, thermalinsulation and acoustic dampening [18, 20, 30].With increasing public demand for clean air, the capture of harmful aerosol particlesby filtration has become the most common method for air cleaning [31]. Fibrous filterswith mean pore size ranging between 100 nm to 100 µm are effective for capturingairborne particles and can obtain high separation efficiencies with a relatively low pressuredrop [31, 32]. A great deal of research has been done towards making efficient filtersfrom pulp in order to find a low-cost and green alternative for polymeric filters [33–35]. Jahangiri et al. [18, 20] studied the effect of fibre morphology and air content offoam-paper on the filtration efficiency and permeability of the product. They generatedaerosol particles by a 1% aqueous solution of Sodium Chloride (NaCl). The aerodynamicdiameter of the particles that were used for filtration tests were between 14 nm to 670 nm.The results showed that permeability of all samples increases by increasing the air contentof the foam. However, by increasing the ratio of refined fibres, permeability of samplesdecreases to a great extent. This happens due to shorter length and higher specific surfacefibres which can block the fluid flow. The results on filtration efficiency of foam-papersshowed that by increasing the air content, the filtration efficiency of foam-paper decreases61.2. Backgrounduntil it loses its filtration properties at 45% air content. However, they found that byadding 10% to 30% of refined fibres in the structure of foam-paper, the filtration efficiencyof the product dramatically increases and the results are comparable with commercialfilters. The effect of adding nanofibrillated fibres (NLF) on the filtration efficiency of theproduct has been studied [18, 20]. The foam made of NLF by freeze drying showed avery high filtration efficiency at high air contents, however it displayed poor mechanicalproperties.Another potential application of foam-papers is using them as thermal insulationmaterials. The effect of porosity and air content on the thermal conductivity of the foam-papers has been studied by Jahangiri et al. [18, 20]. The results showed that a slightchange in porosity has a great impact on the values of thermal conductivity. By increasingthe porosity of the samples, the volume fraction of air increases and consequently thethermal conductivity decreases. Comparing the results with commercial heat insulatorsshowed that highly porous foam-papers could provide a competitive alternative.Porous materials are generally used as sound absorbing materials due to their ef-ficiency in attenuating acoustic energy. Many studies have been done on the soundabsorption by the porous materials [36–39]. Jahangiri et al. [18] studied the effect ofvarious parameters such as air content, thickness, porosity and consistency of the foam-paper. The results showed that the absorption coefficient decreases by increasing theair content of the foam-paper. Increasing the thickness and consistency of the productresults in increasing the acoustic dampening of the foam-paper. The comparison betweenfoam-papers and commercial acoustic products showed that these materials can be ap-plied as sound absorbing materials. The advantages of using foam-papers instead of someother commercial products are that they are degradable, lower cost, and have a reducedenvironmental footprint.1.2.2 Mycelium and sustainabilitySince the industrial revolution, humans have had a widespread impact on the planetthrough industrial wastes and pollutants. In addition to the grave environmental impact,this has also led to serious health concerns of people exposed to these pollutants. The71.2. Backgroundproblems are serious, but fortunately nature has provided us with a potential sustainablebiological solution. Fungi are the recyclers of our planet. Their ability to disassemblecomplex organic molecules into simpler forms helps ecosystems to regenerate. Fungiabsorb nutrients and break down biological polymers through their mycelium. Mycelium,as shown in Figure 1.2, is the subterranean mass of string-like hyphae that functions asfungal roots by absorbing nutrients. Through the mycelium, a fungus absorbs nutrientsfrom its environment by a two stage process: first, hyphae secret enzymes into the foodsource which break down large organic molecules to smaller units such as monomers;then, these monomers are absorbed into the mycelium [40, 41]. That is why it is believedmycelium is the neurological network of nature which has a unique ability to break downand detoxify a great deal of toxic industrial waste and pollution.Figure 1.2: Image of mycelial strands.MycoremediationClean technologies focus on the use of biological methods for the remediation of waste.The process of using fungi to degrade or eliminate contaminants present in water, soils,or air is known as mycoremediation which is a form of bioremediation. This naturaldegradation process is superior to physical or chemical processes because it is less expen-sive and causes minimal site disruption. Fungi use three different methods to degrade81.2. Backgrounda great deal of environmental pollutants: biodegradation, biosorption, and bioconver-sion [42]. Biodegradation is the process which leads to complete mineralization of thestarting compound to simpler ones by living organisms. Many studies have been doneon the degradation abilities of mycelium as represented in Table 1.1.Table 1.1: Role of mycelium in degradation of pollutants.Mushroom spp. Waste/Pollutants RemarkPleurotus ostreatusDDT(dichlorodiphenyl-trichloroethane)DDT was degraded by 48%during a 28 d incubationand 5.1% of the DDT wasmineralized during a 56 dincubation[43].Pleurotus ostreatusPAHs (polycyclic aro-matic hydrocarbons)Suitable for degradationof PAH-contaminatedoil-based drill cuttings[44].Pleurotus sajor-caju PAHsFungal mycelia of Pleurotussajor-caju degraded 99.99%of PAH-contaminatedsoil[45].Pleurotus ostreatusOxytetracycline(OTC)The fungus completely de-grades the drug in fewdays[46].Pleurotus ostreatus Crude oilTotal hydrocarbon concen-tration were reduced by85% [47].Pleurotus ostreatusOxo-BiodegradablePlasticThe plastic was degradedwithout prior physicaltreatment[48].Biosorption is another important process of removal of heavy metals from the en-vironment. The passive uptake of toxicants/metallic ions/xenobiotics by live or driedbiomass can be defined as biosorption. The removal of heavy metal from wastewater isnow shifting from the use of conventional methods to the use of biosorption [49]. Manystudies have been done on the biosorption capacity of mycelium which are summarizedin Table BackgroundTable 1.2: Role of mycelium in the removal of pollutants by using biosorption process.Mushroom spp. Waste/Pollutants RemarkPleurotus ostreatus CadmiumMechanism of biosorptionwas observed[50].Pleurotus mutilus Metribuzin pesticideAdsorption rate ofMetribuzin of 70% wereobtained at optimumconditions[51].TrichodermaNickel, Cadmium, andChromium ionsThe dried biomass of Tri-choderma can be used forthe treatment of toxic heavymetal ions from the indus-trial effluents [52].Pleurotus ostreatus Zinc ionsPleurotus ostreatus is a po-tential adsorbent in wastew-ater treatment due to itsgreat sorption capacity andlow cost[53].Pleurotus sapidus Uranium(VI) ionsPleurotus sapidus is a po-tential adsorbent for Ura-nium(VI) ions removal[54].Bioconversion is a process of conversion of industrial sludges into some other usefulforms. Mushroom is the most important bioconversion product. Any lignocellulosicwaste, generated from industries, can be used for cultivation of mushroom. Kulshreshthaet al. [55, 56] successfully cultivated Pleurotus citrinopileatus and Pleurotus florida onthe sludge of handmade paper and cardboard industrial waste. Other studies on thecultivation of mushrooms from industrial wastes or residues (including cotton waste, ricestraw, sugar beet pulp, and etc.) reported the greatest degradation of the componentsof the substrates as a result of successful cultivation of different mushrooms [57, 58].101.2. BackgroundMycofiltrationAnother application of mycelium is to use it as a membrane for filtering out microorgan-isms, pollutants, and silt. The process of using fungal mycelia to filter toxic waste andmicroorganisms from the environment is called mycofiltration. Mycofiltration membranescan be applied in farms, urban areas, factories, roads, and harmed habitats [59]. Thesupports mostly used in biofilters are organic materials such as soil, compost, peat, bark,due to their large availability and low cost [60]. One of the problems associated withfungal growth is heavy mycelial growth occupy the void space in filters which results inincreased pressure drop and clogging [61, 62]. This problem causes a significant decreasein the elimination capacity of membrane. Aizpuru et al. [63] used porous ceramic ringsto control pressure drop. The study also showed that fungal biofilters have a greater po-tential to remove toluene vapor as compared to bacterial systems. Another applicationof mycofilters is pathogen management. Taylor et al. [64] developed a mycofilter whichis capable of filtering E. coli from stormwater.1.2.3 Filter mediaA filter medium is any material that, under the operating conditions of the filter, ispermeable to one or more components of a mixture, solution, or suspension and is imper-meable to the remaining components [65]. In the filtration process, the surface strainingis when the particle is too big to enter the entrance of the pore, but the particles mayfit through the entrance and may be trapped by one of the four mechanisms of particlecapture after they penetrate the surface of the medium [66], as listed below:• Inertial impaction − occurs when the particle inertia is so high that it has sufficientmomentum to break away from the streamlines and adhere to the wall of the pore.• Interception − occurs when small particles do not have enough inertia to breakdown the streamlines, however the streamlines may carry them too close to theobstructions where surface forces cause the particles to adhere to the fibres.• Diffusion − occurs when very small particles (<0.5 µm) exhibit Brownian motion111.2. Backgroundand move randomly due to collisions with other particles. They may randomlytouch the wall surface of the pore.• Electrostatic attraction − occurs due to electrostatic charge on the particle and/orfibre that will force the particle to be attracted to the fibre with an opposite charge.Both inertial impaction and Brownian diffusion are much less effective with liquidsthan with gases [66]. Since the density of a particle is closer to the liquid than a gas,deviation of a suspended particle from the liquid is much less, thus the adherence ofa particle by inertial impaction is less likely. Brownian diffusion in liquids occurs to alimited extent because Brownian motion in liquid suspensions is not as noticeable as ingaseous suspensions.Fibrous materials are a low cost filter media, the fibres can be derived from natural orsynthetic sources [65]. Fibrous filters are widely used in many industries, such as phar-maceutical, biotechnology, microelectronics, and semiconductor manufacturing. Fibrousfilter media can be classified as woven and nonwoven filters, where nonwoven filters aremore commonly used in filtration technology. The properties of fibrous filters is relatedto the properties of the fibres. Finer fibres are able to trap smaller particles and improvethe filtration performance. However, finer fibres produce a weaker filter media. To findthe best filter for a specific application, filtration efficiency, pressure drop and mechanicalstrength must be optimized by selecting the appropriate fibres. A weak material withgood filtration performance can be strengthened by supporting it on a stronger substratebut, this often carries an associated manufacturing cost increase which can limit thecommercial viability of the product.Throughout the history of filtration and separation processes, fibrous media in theform of nonwoven filters have been used extensively in water treatment, water desali-nation, and water discharge treatment plants [65]. Compared to asymmetric membranefilters, nonwoven filters have a high internal surface area which leads to an increase thedirt loading capacity of the filters [67]. Despite having advantages of low cost, high dirtholding capacity, and high filtration efficiency, nonwoven media is limited to removingparticles with diameter between 10 and 200 microns [68].121.3. Cellulose-Mycelia Foam as a filtration media1.3 Cellulose-Mycelia Foam as a filtration mediaIn pursuit of sustainable products that leverage Canada’s available natural resources,this research is conducted to produce a novel eco-friendly biocomposite material, whichis called cellulose-mycelia foam (CMF). In this product, cellulose fibres are used as anatural reinforcement, and mycelium act as a bio-based matrix to surround and supportthe cellulose fibres.The procedure of making CMF is quite similar to making foam-paper, with the addi-tion of specific fungi grain spawn. In the foam making stage, the air content and consis-tency of foam are kept constant to evaluate the effect of mycelial growth on mechanicalproperties of the CMF biocomposite. The samples are kept at cultivation conditionsup to four weeks. After observing the desired mycelial growth, they are dried in a lowtemperature oven to prevent any further mycelial growth. The final product has 3-Dstructure, high porosity, high strength and is extremely light. This characteristics of theCMF position this biocomposite as a green alternative to polymeric foams. The CMFalso can be used in a wide range of applications including insulation, packaging, filtrationand acoustic damping.In the first experimental study, the morphology, thermal stability, compressibility,energy absorbing, wet and dry tensile strength of the CMF are assessed to characterizethe mechanical properties of the product, presented in Chapter 2. In all experiments, thetime of mycelial growth is considered as an input variable, while other factors includingconsistency, fibre length, diameter, and properties of fibres are constant.In the second experimental study (Chapter 3), an application of the CMF as a hy-draulic filter is studied. Pressure drop, permeability, hydraulic filtration efficiency, andfilter quality factor are measured in a small flow circuit. As mycelia can remediate con-taminate present in water, soils, or air, the effect of mycelia on the mycoremediationabilities of the CMF to degrade potassium hydroxide are investigated in final part of thestudy.13Chapter 2Cellulose-Mycelia Foam: productionand characteristics2.1 ProductionOften the limiting factor in adopting foam-paper for an application is strength. Toachieve a highly porous foam-paper, low consistency and high air content is required;although these lead to desirable increases in porosity, it also adds weakness in the finalproduct. One way of circumventing this problem is to increase the weight percentage ofrefined fibres in the structure of foam-papers. Another approach to solve this problemis to add mycelium to the final product. Since mycelium spreads out through the sam-ples, effectively reinforcing the material, it is a good candidate to make a high porous,biodegradable, and sustainable biocomposite with improved mechanical properties. Fig-ure 2.1 shows a sample of the cellulose-mycelia foam.(a) Top view (b) Side viewFigure 2.1: Image of dried cellulose-mycelia foam after 25 days.142.1. ProductionIn this study, the methodology for production of cellulose-mycelia foam (CMF) isquite similar to making foam-paper [18], with the addition of specific fungi grain spawn.A standard Northern Bleached Softwood Kraft (NBSK) pulp sheets were used as the fibresource for making 1% consistency pulp/water suspension. In this process, a Bioterge AS40 surfactant was added to NBSK pulp/water suspension in 10 wt% at pH of 7, asa foaming agent. Second, the suspension of foam and fibres was rapidly agitated toreach 50% air content. Third, the mushroom grain spawn and nutrients (sawdust andcornmeal) were added to the foam suspension while the mixture was being agitated. Thesuspension was filtered in a 10 cm diameter Buchner funnel under 9.8 kPa of vacuum.Then, the filtered foam was kept at cultivation conditions for 5 to 25 days. Finally, afterthe desired growth of mycelium was observed, the foam was dried in a low temperatureoven at 55 ◦C for 2 hours, to inhibit any further mycelium growth. The characteristicsof the samples that were used for testings are briefly reported in Table 2.1.Table 2.1: Characteristics of the cellulose-mycelia foam samples.Mushroom ssp. Pleurotus DjamorPulp type NBSKConsistency 1 %Air Content 50 %Growth time 5 - 25 daysDrying method Oven-driedDrying temperature 55 ◦CDrying time 2 hoursFungi grain spawn is used to introduce the fungi into the paper media. Commerciallyavailable prepared fungi grain spawn was used. A certified organic rye grain spawn wasused which is ideal for inoculating the sterilized substrate, or for use in unsterilizedstraw, paper or bed-style inoculations. In this study, Pleurotus Djamor (Pink OysterMushroom) was cultivated because it grows faster than other species. Each species of thefungi requires specific cultivation conditions. In general, fungi do not have chlorophylland do not perform photosynthesis, therefore exposure to sunlight is not mandatory.However, it does not mean fungi necessarily require a dark environment to grow. The152.1. Productionadvantage of growing in the darkness is that dark areas often provide the moisture thatthe spores need to reproduce. Because fungi are not able to retain the moisture, anenvironment that has a high humidity is mandatory to avoid water loss. Based ondifferent studies [69–74] on mycelium growth especially on the Pleurotus species, thebest cultivation conditions are listed below:• Temperature: 20 - 25 ◦C [69–71].• pH: 5 - 8 [69].• Humidity: 80 - 100%.• Light: darkness.• Ventilation: mushrooms breathe and exchange gases, so air circulation and gasexchange is required.• Time: generally, between 2 to 4 weeks are ideal for mycelium running [70–72].• Nutrients: cellulose, lignin, fibre content of substrate, husk rice, straw, and corn.Based on a study on the performance of Pleurotus ostreatus [72], total average ofmycelium growth time for Pleurotus species is 15 to 25 days. Another study on cultivationof Pleurotus species [71] showed that the complete spawn run is 2, 3, and 4 weeks forpaper, fibre and sawdust respectively. Baysal et al. [70] found that the addition ofhusk rice to waste paper increases the mycelial growth. Another study on the growthof Pleurotus species [73] showed that the yield of mushroom is positively correlated tocellulose, lignin, and fibre content of substrates. In this study, straw and cornmeal wereused as excess nutrients to improve mycelial growth. Park et al. [69] found that theoptimal corn steep powder concentration is 10 g/L. Therefore, 1 wt% of nutrients wereadded to the pulp suspension.162.2. Morphology and thermo-mechanical properties2.2 Morphology and thermo-mechanical propertiesOne of the major problems associated with mycelial growth is that mycelia tend tooccupy the void space between fibres. It is necessary to study the morphology of theproduct to find the optimum growth time based on the intended use of the final product.In this section, the morphological characteristics of the CMF during mycelial runningwere studied. In this research, the time of mycelial growth is considered as an inputvariable, while metrics such as mass loss, compressibility, and tensile index (TI) are usedto evaluate the effect of growth time on the final product.2.2.1 MorphologyFungi are morphologically complex microorganisms, exhibiting different structural formsthroughout their life cycles [75]. The basic vegetative structure of growth consists ofa tubular filament known as hypha that originates from the germination of a singlereproductive spore. As the hypha continues to grow, it frequently branches repeatedly toform a mass of hyphal filaments referred to as mycelium. Mycelia secrete a wide varietyof enzymes into the environment [76]. Most enzymes are secreted at the edge and inthe centre of mycelium to aid in digestion of surrounding organic material. In proteins,and therefore enzymes, the indole group of tryptophan display intrinsic fluorescence [77].Intrinsic fluorescence has become an important evaluation tool in various biologicallyrelevant studies including morphological studies of mycelium [78–80].The morphology of the CMF during the growth time of mycelia was studied by usinga Nikon Eclipse TE200 fluorescent microscope. A filter cube with 520 - 540 nm excitationand 560 - 620 nm emission was used to attempt to excite and measure the fluorescenceof the indole group present in the mycelial enzymes. Figure 2.2 shows the images of thecross-section of CMF.Fluorescent microscopy revealed that mycelia grow and cover the cellulose fibres.Figure 2.2a showed that the cellulose fibres in the structure of foam-paper (0 days CMF)did not display intrinsic fluorescence, and the fibres were dark in the absence of mycelia.After 10 days of mycelial growth, the fibres became brighter because of mycelial runningwhich leads to the secretion of enzymes (Figure 2.2b). The results showed the cellulose172.2. Morphology and thermo-mechanical properties(a) 0 days CMF (b) 10 days CMF (c) 25 days CMF(d) 0 days CMF (e) 10 days CMF (f) 25 days CMFFigure 2.2: Fluorescent micrographs (top) and conventional photographs (bottom) show-ing the effect of mycelial growth on the microstructure of cellulose-mycelia foam. Note:field of view for all micrographs is 1.4 mm x 1.6 mm.fibres were covered by mycelial strands after 25 days, as shown in Figure 2.2c. Also,assuming that the excreted enzymes are the primary source of fluorescence, it can beconcluded that secretion occurred more in central zones of the colonies compared to theperiphery of the mycelia.As mycelia grows, the volume fraction of cellulose decreases due to the degradationof the cellulose fibres by mycelia as a nutrient and mycelial strands replace the cellulosefibres and occupy the void spaces between them. However, decreased volume fraction ofcellulose is negligible compare to increased volume fraction of mycelia in the first 25 days,as extra nutrients are provided in the substrate, and the growth rate is fast. Thus thevoid spaces between fibres are occupied by mycelial strands which can lead to decreased182.2. Morphology and thermo-mechanical propertiesporosity of the biocomposite, as can be expressed by the following equation:ff + fm + fv = 1 (2.1)As mycelia grows, the summation of the volume fraction of cellulose fibres ff and volumefraction of mycelial strands fm increases, and the void fraction fv decreases, which leadsto decreased porosity and a densely packed structure of the CMF.2.2.2 Thermal Gravimetric AnalysisTo study the effect of mycelial growth on the thermal stability of the CMF, the amount ofweight loss as a function of increasing temperature was obtained by thermal gravimetricanalysis (TGA), as shown in Figure 2.3. Samples were run in a nitrogen atmosphere ona Q500 TGA manufactured by TA Instruments. The samples were heated from 30 ◦C to600 ◦C, with a constant purge gas rate of 40 ml/min and a scanning rate of 10 ◦C/min.0 200 400 600Temperature (°C)020406080100Normalized mass remaining (%)0 days15 days25 daysFigure 2.3: The effect of mycelial growth on the thermal stability of the cellulose-myceliafoam.192.2. Morphology and thermo-mechanical propertiesThere are three stages of degradation in the TGA curves of all samples.The initialmass loss below 120 ◦C can be attributed to the evaporation of moisture. This lossdepends on the initial moisture content of the fibres. The second, severe weight loss(250 − 350 ◦C) and third stage are due to decomposition of the major components ofthe fibres. The TGA curve of the NBSK cellulose foam, labelled as 0 days in Figure 2.3,showed the thermal decomposition of cellulose begins around 227.5 ◦C (extrapolatedonset temperature); while, the temperature at which the weight loss begins increaseswith the presence of mycelia. The extrapolated onset temperatures are 312.7 ◦C and325.5 ◦C for 15 days and 25 days respectively. After 600 ◦C, only ash and char were left,0 100 200 300 400 500 600Temperature (°C)050100Normalized mass remaining (%)-10Derivative weight (%/°C)(a)0 100 200 300 400 500 600Temperature (°C)020406080100Normalized mass remaining (%)-2-1.5-1-0.500.5Darivative weight (%/°C)(b)0 100 200 300 400 500 600Temperature (°C)020406080100Normalized mass remaining (%)-2-1.5-1-0.500.5Derivative weight (%/°C)(c)Figure 2.4: TGA curve and first derivative of the weight loss for (a) 0 days (b) 15 daysand (c) 25 days CMF.202.2. Morphology and thermo-mechanical propertiesand the remaining mass was 10% for all samples. It can be concluded that mycelialgrowth improves the thermal stability of the final product. The best result was obtainedafter 25 days of mycelial growth which delayed the onset of thermal decomposition tohigher temperatures.Another significant characteristic is the point of greatest rate of change on the weightloss curve. This is also known as the inflection point. The peak calculation of the 1stderivative of the weight loss curve indicates the inflection point of the sample, as shownin Figure 2.4. The noise in the derivative plots is due to random noise in the originaldata. The inflection point temperature was calculated to be 286.9 ◦C, 342.4 ◦C, and360.3 ◦C for 0 days, 15 days, and 25 days of mycelial growth time respectively. Theresults indicated the same trend as extrapolated onset temperature, with the best resultsobtained after 25 days of mycelial growth. It showed the greatest rate of change occursaround 360 ◦C which is much greater than the temperature for the pure foam-paper.2.2.3 Dynamic Mechanical AnalysisTo study the effect of mycelial growth on the compressibility of the CMF, the stress-straincurve was obtained by dynamic mechanical analysis (DMA). The test was performedusing a DMA Q800 manufactured by TA Instruments operating in controlled force mode.The samples were cut into cylinders of diameter D = 15 mm by thickness L0 = 10 mm.The samples were placed in compression clamps under isothermal conditions at 22 ◦C,and the compressive force was ramped at a rate of 0.5 N/min up to a maximum of 2.0 N.Stress σ and strain ε are recorded throughout the test, and are defined as follows:σ =4FpiD2(2.2)ε =xL0(2.3)where F is measured compressive load, x is displacement which is measured from theinitial dimension of the sample.The compressive stress-strain curve of samples with different growth days are pre-212.2. Morphology and thermo-mechanical properties0 20 40 60 80Absolute Strain (%)02468Stress (MPa)#10-30 days15 days25 daysDensification stageLinear elastic stagePlateau stageFigure 2.5: The effect of mycelial growth on the compressibility of the cellulose-myceliafoam.sented in Figure 2.5. Typical of other porous materials, the compressive stress-straincurve indicates three distinct stages: the linear elastic stage, the plateau stage, and thedensification stage. At low strains (<5%), the stress-strain behaviour is linear elasticin which the stress elevates linearly with increasing strain. The compressive test resultsshow that in the linear elastic region, the compressive modulus of elasticity is higher forboth CMF samples as compared to the foam-paper sample.In the plateau stage, the stress increases slowly in a wide strain range, which meansthe structure begins to collapse at an approximately constant stress. The foam-papershowed higher deformation at the constant stress which is due to its high porosity compareto the CMF samples, as shown in Figure 2.5. The 15 days and 25 days CMFs which havemore compact structure, resulting in an increase in the plateau stress and a significantreduction in the densification initial strain.222.2. Morphology and thermo-mechanical propertiesIn the third region of the compressive stress-strain curve, referred to as the densifica-tion region, the complete collapse occurs with the characteristics of a rapid increase ofstress with increasing strain. The densification initial strain was calculated to be 55%,30%, and 20% for foam-paper, 15 days and 25 days CMF respectively.The area under stress-strain curve can be related to the energy absorbtion of thematerial [81]. To compare the effect of mycelial growth on the energy absorption charac-teristics, the work of compression Wcomp was calculated for each sample. The compressivework is related to stress σ and displacement x as follows:Wcomp =∫Fdx =pi4D2∫σdx (2.4)Test results were recorded at discrete intervals, so the compressive work is approxi-mated by numerically integrating Equation 2.4 as follows:Wcomp ≈ pi4D2L0N∑iσi(εi+1 − εi) (2.5)The compressive work is 6.16 mJ, 171 mJ, and 90 mJ for 0, 15, and 25 days respec-tively. Compared with foam-paper, the both CMF samples revealed at least 10 timeslarger compressive work which indicates better energy absorbing properties. It can beconcluded from the aforementioned results that growing mycelium in the structure of thefoam-paper can improve the compressive strength and energy absorbing characteristicsof the biocomposite. It is hypothesized that mycelia grow through the foam paper pro-viding additional structure to the material – acting as a matrix. This leads to decreasedporosity of the biocomposite and improved compressive strength.2.2.4 Tensile and wet strengthMany factors should be taken into consideration when designing a biocomposite, includingthe length, diameter, orientation, consistency, and properties of fibres; the properties ofmatrix; and the bonding between the fibres and the matrix [81]. The effect of length,diameter, and consistency of fibres on the structure of foam-paper have been discussedin Section 1.2.1. To study the effect of mycelial growth on the strength of the CMF as232.2. Morphology and thermo-mechanical propertiesa matrix, the tensile strength test was performed during mycelial running by Thwing-Albert’s QC Electronic Materials Tensile Tester. The gauge length and the speed werefixed at 1 cm and 2.0 cm/min respectively. The samples were cut into 15 mm widestrips with sufficient length to be clamped in the jaws as described in TAPPI T 494.The samples were tested in accordance with the standards outlined in TAPPI T 494 andT 456. The dry and wet tensile index were calculated by using the following equations:F =vL(2.6)TI =FwR(2.7)where F is the breaking force, v is the separation speed of the jaws, L is the readingload, TI is the tensile index in Nm/g, w is the specimen width in meters, and R is thegrammage in mass per unit area (g/m2).0 5 10 15 20 250123Growth time (day)Tensile index (N.m/g)  Dry CMFWet CMFFigure 2.6: The effect of mycelial growth on the tensile index and wet strength of thecellulose-mycelia foam.242.2. Morphology and thermo-mechanical propertiesFigure 2.6 shows the effect of mycelial growth on the dry and wet strength of theCMF. The results revealed that by increasing the growth time of mycelia in the foam-paper structure, wet and dry tensile index increases at a constant bulk. The dry tensileindex of the CMF improved from 0.61 Nm/g for foam-paper to 2.65 Nm/g for 25 daysCMF. The wet strength also showed the same trend. The wet tensile index increasedfrom 0.31 Nm/g for foam-paper to 1.38 Nm/g for 25 days CMF.As we discussed in Section 2.2.1, the mycelial strands occupy the spaces betweenfibres during incubation period and decrease the porosity of the CMF, which can lead tohigher tensile strength at the constant bulk. Another explanation for increased strengthcan be expressed by the relation of the stress taken by the fibres, the stress taken by thematrix and the strength of the composite [81]. The strength of the composite σc can beestimated fromσc = ffTSf (1− lc2l) + fmσm (2.8)where ff and fm are the volume fraction of fibres and matrix respectively, TSf is thetensile strength of the fibre, lc is the critical fibre length, and σm is the stress on thematrix at failure. A critical fibre length for any given fibre diameter can be determined:lc =TSfd2τi(2.9)where d is the diameter of the fibre, τi is related to the strength of the bond betweenthe fibre and the matrix [81]. Substituting Equation 2.9 into Equation 2.8 yields anexpression for the strength of composite in terms of the contribution from the fibres,matrix and the bond strength between them:σc︸︷︷︸Strength ofcomposite= ffTSf︸ ︷︷ ︸Stress takenby fibre+ fmσm︸ ︷︷ ︸Stress takenby matrix− ffTS2fd4τil︸ ︷︷ ︸Bond betweenfibre & matrix(2.10)Equation 2.10 shows that the strength of composite is proportional to the stress takenby fibre, the stress taken by matrix, and the bond between them. Usually the stress that252.2. Morphology and thermo-mechanical propertiesσfσfσcσca) b)Cellulose fibresMycelial hyphaFigure 2.7: a) Interaction between cellulose fibres in foam-paper, b) interaction betweencellulose fibres and mycelia in the cellulose-mycelia foam composite.the matrix can hold until it fails σm, is much smaller than tensile strength of the fibreTSf . Hence, the larger the fraction of matrix fm is, the smaller the strength of thecomposite would be if the other factors are constant. The only term left in Equation 2.10which effects the strength of the composite is the bond between the fibres and matrix.With mycelial growth, the strength of the bond between cellulose fibres and the mycelialstrands τi increases. As shown in Figure 2.7, the mycelial filament network attach tomultiple cellulose fibres improving the connection between individual fibres. This resultsin reduced critical fibre length lc. It can be concluded that the tensile strength of theCMF can be optimized by controlling mycelial growth.26Chapter 3Applications3.1 Water filtrationFibrous media in the form of nonwoven filters are widely used in processing water andwastewater as pre-filters [82]. We consider the suitability of the CMF as a biocompositefilter media to eliminate undesirable contaminants from a water system. In this section,the filtration properties of the CMF are studied. The effect of mycelial growth on thepressure drop, permeability, filtration efficiency, and filter quality factor of the CMF areevaluated.3.1.1 BackgroundFoam-papers have a very high porosity, high dry filtration efficiency, and a low resistanceto flow [18]. If the foam-paper wets, the fibres will absorb water which leads to twosignificant consequences: the fibres swell, so the void spaces between them reduce whichcan improve the filtration efficiency, however the mechanical strength drops sharply. Tobe used as a wet filter, the foam must be fully supported by a strong substrate. Oneof the original purposes of introducing cellulose-mycelia biocomposite was to providemechanical support for the foam for liquid filtration. The effect of mycelial growth onthe wet strength of the CMF is shown in Figure 2.6, which shows a significant increasein the wet strength with increasing mycelial growth.The CMF is a wet laid nonwoven filter media with its porous fabric composed of arandom array of mycelial fibres within the cellulose fibres. Liquid filtration of nonwovenfilter media can not be the only method of purification, because particles of 1 µm orless size can pass through, and further separation is required [66]. It’s generally assumedthat the flow through nonwoven filter media, which is dominated by viscous effects at273.1. Water filtrationlow Reynolds numbers, can be described by Stoke’s law. In the creeping flow regime, theeffect of gravity and inertia become negligible compared to viscous forces, and Darcy’slaw can be applied to nonwoven filter medium used in liquid filtration. Darcy’s law givesa general description of fluid velocity as a function of pressure gradient, fluid viscosity,and hydraulic permeability. Darcy’s law and calculating hydraulic permeability will bediscussed further in Section MethodsInvestigations on the performance of the CMF were conducted for the foam-paper (0 days),5, 15, and 25 days CMF. The filter pressure drop of samples were measured in a smallcircuit as shown in Figure 3.1. Each CMF sample was cut into 45 mm diameter disks tofit into the sample-holder in the circuit. Water was pumped into the circuit, and the flowrate was controlled by a low-flow flowmeter with a range of 2 to 22 gph. The pressuredrop of the samples were measured by a differential pressure sensor measuring across theinlet and outlet of the sample holder.		Flow	Controller		Pump		Pressure	Transducer	Sample	Holder		Water	Supply	Figure 3.1: The hydraulic circuit for measuring the pressure drop of the CMF filters.To evaluate the filter performance of the CMF, testing was conducted in accordance283.1. Water filtrationwith ISO 12103-1, A2 FINE TEST DUST [83]. Particles varying in diameter from1 µm to 45 µm were added to the fluid until it reached a specified concentration ingrams of dust per litre of water. This solution was circulated continuously at 5 gph(Q = 5.26× 10−6 m3/s). The performance of the filters were determined using the Mas-tersizer 2000 (Malvern, UK) by measuring the particle size distribution of the upstreamand downstream flows. The measurements were conducted using a Hydro MU sampledispersion unit, and applying 20 s of ultrasound treatment. In this study, the Mie theorywas applied, assuming the following values for the indices: particle refractive index 1.572,particle absorption index 0.1, and refractive index 1.33 for water as a dispersant.3.1.3 Results and discussionFigure 3.2 shows the experimental values of the pressure drop against flow rate for var-ious mycelial growth days under continuous loading of water. The results revealed thatincreasing the growth days of mycelia in the structure of the foam leads to an increasedpressure drop(∆P) at a constant flow rate. The resistance to fluid flow through the CMFis related to mycelial growth that occupies the void spaces between the cellulose fibreswhich leads to a decrease in the porosity of the final product.The permeability is a constant intrinsic to the porous medium. The permeability ofthe filters were calculated by using Darcy’s law which describes the flow of a fluid througha porous medium. For a finite 1-D flow as shown in Figure 3.3, Darcy’s law is stated asq =QA(3.1)q = k∆PµL(3.2)where Q is the volumetric flow rate (m3/s), A is cross-sectional area (m2), q is thevolumetric flux (m/s), ∆P is the pressure gradient (Pa), L is the flow path length (m),µ is the fluid viscosity (Pa · s), and k is the hydraulic permeability (m2). In Figure 3.4the normalized permeability is shown as a function of mycelial growth days. The resultsshowed that the permeability of the CMF decreases by increasing the mycelial growth293.1. Water filtration0 1000 2000 3000 4000 50000481216x 10−6∆P (Pa)Q (m3 /s)  0   days5   days15 days25 daysFigure 3.2: The effect of mycelial growth on the pressure drop of the cellulose-myceliafoam under continuous loading of water.days. An increase in the growth days of mycelia leads to a reduction in the porosity ofthe final products which diminishes the hydraulic permeability.	L A Q Dp PT Figure 3.3: Schematic view of a 1-D model of the filtration apparatus used to calculatethe permeability of the CMF samples.303.1. Water filtration0 5 15 25Growth time (days) / df2Figure 3.4: The effect of mycelial growth on the normalized hydraulic permeability ofthe cellulose-mycelia foam.Filter performance is often expressed in terms of percent efficiency, defined as theratio of upstream particle concentration compared to the downstream concentration thathas passed through. Figure 3.5 shows the hydraulic filtration efficiency of the CMF as afunction of particle diameter for different mycelial growth time. An increase in mycelialgrowth time leads to an increase in filtration efficiency of the product for all particlediameters. The filtration efficiency for foam-paper (0 days) was the lowest and that for25 days CMF was the highest. This is because the 25 days CMF is lower in porositycompare to the others and interception is the dominant mechanism in hydraulic filtration.It is more likely that the particles touch the surface of the pore wall in the 25 days CMF.The most commonly used rating in industry to evaluate the hydraulic filter perfor-mance is the Beta ratio. Beta ratio is defined as the ratio of the number of particlesupstream versus the number of downstream, greater than a given size. Table 3.1 showsthe values of Beta ratio and capture efficiency of the CMF filters for 5 µm, 10 µm, and20 µm. The results showed that the foam-paper (0 days) is 17% efficient at removing313.1. Water filtration100 101 102Particle diameter (7m)020406080100Efficiency (%)0 days5 days15 days25 daysFigure 3.5: The effect of mycelial growth on the hydraulic filtration efficiency of thecellulose-mycelia foam.5 µm and larger particles, and 74% efficient at removing 20 µm and larger particles,while the 25 days CMF is 50% and 99.9% efficient respectively. The experimental resultsclearly demonstrated that mycelial growth improves the hydraulic filtration efficiency ofthe CMF.Table 3.1: Selected Beta ratio and the corresponding efficiencies of the CMF filters for5 µm, 10 µm, and 20 µm.Filtertypeβ(5)Captureefficiency(%)β(10)Captureefficiency(%)β(20)Captureefficiency(%)0 days 1.2 17 1.5 34 3.8 745 days 1.25 20 1.7 41 5 8015 days 1.5 35 2.1 54 6.7 8525 days 2 50 5 80 1000 99.9323.1. Water filtrationFilter quality factor is another useful parameter to characterize filter performancewhich takes both the effect of efficiency and pressure drop into account. Filter qualityfactor is calculated asQF =−ln(1− η)∆P(3.3)where η is the filtration efficiency, and ∆P is the pressure drop across the filter (Pa).Figure 3.6 shows the calculated quality factor of the CMF for various particle diametersat a constant flow rate (Q = 5.26× 10−6 m3/s). The results shows that 15 days of CMFis more efficient than other samples for smaller particles, while 25 days CMF is a betteroption for larger particles.100 101Particle diameter (μm)00.511.522.53Quality factor (1/Pa)10-30 days5 days15 days25 days1.5 2 2.5 3 3.5 4 4.5 511.522.533.54 10-4 × ×Figure 3.6: Quality factor of the cellulose-mycelia foam as a hydraulic filter media.333.2. Mycoremediation3.2 MycoremediationThe use of fungi to reduce or eliminate contaminate present in water, soils, or air isknown as mycoremediation. Pleurotus species are a widely cultivated mushroom dueto their strong ability to degrade a wide variety of materials and compounds [84]. Inthis section, the effect of dead and living mycelia on the mycoremediation abilities ofP. djamor mycelia in the cellulose-mycelia foam to degrade KOH were investigated.3.2.1 BackgroundA battery is an electrochemical device which consists of an anode, a cathode, an elec-trolyte, separators, and the external case. The materials used as electrodes and elec-trolyte are the main differences between different battery systems. Separators are madeof polymeric materials, paper, or paperboard. The external case is composed of steel,polymeric materials or paperboard. The consumption of batteries has increased becauseof low maintenance, reduced cost, and its requirement by the electronic industry [85].Based on statistics for Canadian battery sales, primary batteries make up approximately78% of total sales, by weight, in the country [86]. Alkaline battery is a type of primarybattery which is the most common battery chemistry in Canada. It makes up 58% ofthe total battery market in Canada [86], as shown in Figure 3.7. Today, the majority ofthese batteries go to landfills at the end of their life cycle. An interest in environmentalissues related to battery disposal has been growing [85, 87].In alkaline batteries, the negative electrode is zinc and the positive electrode is man-ganese dioxide, and the electrolyte is potassium hydroxide (KOH). The cell is totallyenclosed in a high density steel, and a separator is made from non-woven fabric to sep-arate the anode and cathode from the electrolyte solution. A typical initial compositionof an alkaline battery is presented in Table 3.2 [88].The zinc and manganese dioxide are consumed during discharge, while the alkalinepotassium hydroxide is not part of the reaction and remains in the batteries. Potassiumhydroxide is colourless, odourless, and corrosive. It is soluble in water up to 1 M at20 ◦C. This hazardous alkaline may leak by increased duration of residence in landfills.One of the most feasible solutions to potassium hydroxide contamination is to use a343.2. MycoremediationAlkaline58%Zinc	Carbon18%Zinc	Air	Button0.1%Lithium	(P)2%Silver	Oxide	Button0.1%Other	Primary	Batteries0.02%NiCd14%NiMH4%Li-ion1%SSLA2%Figure 3.7: Market share for batteries, based on tonnes sold in Canada by KelleherEnvironmental [86].bioremediation process. This study evaluates the ability of the CMF samples to detoxifypotassium hydroxide from discharged alkaline batteries.Table 3.2: An average composition of Alkaline battery [88].Battery component Typical composition (%)Manganese dioxide 32− 38Zinc 11− 16Potassium hydroxide 5− 9Graphite 3− 5Steel 19− 23Barium sulfate <5353.2. Mycoremediation3.2.2 MethodsTo investigate the bioremediation capabilities of the CMF samples, four different sampleswere used: foam-paper, live mycelia after 5 days of growth, live mycelia after 25 days ofgrowth, and dried CMF after 25 days of growth. Before adding live and dead samples,leaking AA alkaline batteries were placed in the water to obtain strong base toxic solution.The equal amount of solution were added to different containers. The pH of all containerswere 12 at the first day of experiment. Then, all different samples were add to thecontainer. A pH meter was used to measure the pH of solutions every 5 days for 60 days.A change in pH indicates the amount of remaining potassium hydroxide in the solution.3.2.3 Results and discussionFigure 3.8 shows the pH of solutions as a function of time. The pH of control samples(foam-paper) showed insignificant changes which can likely be attributed to experimentaluncertainty in the pH measurement. The pH of the solution with dead samples after 25days of incubation decreased slightly for 20 days and then remained the same for another40 days. The pH of the solution for both living mycelia added after 5 days and 25 daysof incubation dropped during the 60 day test period. The results for living mycelia after25 days of incubation showed a sudden decrease in the amount of pH after 20 days.A decrease in the amount of pH shows a reduction in the hydroxide ion concentrationwhich shows potassium hydroxide was used as a nutrient for mycelia by biosorption mech-anism. The following equations define the relation between the pH and the hydroxideion concentration.pOH = 14− pH (3.4)[OH−] = 10−pOH (3.5)It can be concluded the living mycelia have the ability to detoxify the potassiumhydroxide solution by biosorption. Figure 3.9 shows the various biosorption mechanismsaccording to the dependence on the cells metabolism [89]. Veglio et al. [89] demonstrated363.2. Mycoremediation0 10 20 30 40 50 60Number of days678910111213pHFoam-paper5 days25 days25 days- deadFigure 3.8: Detoxification of KOH by dead and live mycelia of P. djamor in contaminatedwater during 60 days.that transport across the cell membrane is associated with cell metabolism and may takeplace only with viable cells. This method of biosorption is not immediate, since it requiresthe time for the reaction of the microorganism. It is hypothesized that mycelia transportthe ions across mycelial cell membranes and convey potassium as an essential mineralnutrient. This is why more mycelial growth leads to faster detoxification as living samplesafter 25 days incubation obtained the best results during the 60 day test period. However,the slight decrease in the pH for dead samples can be attributed to physicochemicalinteraction between the ions and functional groups of the cell surface, based on physicaladsorption, ion exchange, and complexation which are not dependent on the metabolism.Also, precipitation is another possible theory, which is either dependent on the cellularmetabolism or independent of it [89]. Mycelia may excrete compounds which favour theprecipitation process.373.2. MycoremediationBiosorption MechanismsMetabolismdependentNon-metabolismdependentTransport acrossthe cell membranePhysicaladsorptionPrecipitation ComplexationIon exchangeFigure 3.9: Biosorption mechanism according to the dependence on the cell metabolism.38Chapter 4Conclusions and future workSince the industrial evolution, humans have had a widespread impact on the planetthrough industrial wastes and pollutants, and the demand for sustainable products con-tinues to grow. The development of green and biodegradable products is crucial to helpecosystem to regenerate. Cellulose mycelia foams were developed by using foam-formingprocess in paper industry. These green biocomposite consist of mycelial strands withinthe cellulose fibres which have a 3D porous structure. The experimental tests showreasonable results which suggest these biocomposite can be useful in a wide range ofapplications.The morphological study of the CMF showed that mycelial strands replace the cel-lulose fibres by decomposition and occupy the void spaces between fibres which lead toa reduction in the porosity of the product. The effect of mycelial growth on the me-chanical properties of the CMF were studied. The thermal gravimetric analysis showedincreasing mycelial growth days increases the onset temperature of thermal decomposi-tion. The mycelial growth improved the thermal stability of the CMF. The 25 days ofmycelial growth improved the onset temperature by 40% compared to the foam-paper.The strain-stress curve of the CMF was obtained by dynamic mechanical analysis. Anincrease in mycelial growth caused an increase in the plateau stress and a significantreduction in the densification initial strain which leads to better energy absorbing prop-erties. The effect of mycelial growth on the dry and wet strength of the CMF also werestudied. The wet and dry tensile index increases by increasing the growth time of mycelia.The dry tensile index improved from 0.61 Nm/g for foam-paper to 2.65 Nm/g for 25 daysCMF. The wet tensile index also increased from 0.31 Nm/g for foam-paper to 1.38 Nm/gfor 25 days CMF.The hydraulic filtration properties of the CMF were evaluated by using a small flow39Chapter 4. Conclusions and future workcircuit to measure the pressure drop and the particle size distribution of the upstreamand downstream flows. The experimental results showed that the resistance to fluid flowthrough the CMF is related to the mycelial growth time. The results also showed thatan increase in mycelial growth leads to a decrease in the permeability, and an increase infiltration efficiency of the product. The experimental results showed that 25 days CMFis 50%, 80%, and 99.9% efficient at removing particles larger than 5 µm, 10 µm, and20 µm respectively.To investigate the detoxification capabilities of the CMF samples, the pH of leakingalkaline battery in water were measured every 5 days for 60 days. The results showedthat living mycelia is effective to detoxify potassium hydroxide, but the dead mycelia isnot, and the CMF samples are not able to detoxify inorganic compounds.The 3-D, lightweight, highly porous CMF can be applied in a wide range of applica-tions and provide a green alternative to polymeric foams. Due to the time limitation ofthis project, some characteristics of the product were studied. To have a better under-standing of the CMF and its suitability for different applications, other characteristicsof the product can be evaluated in future projects such as sound absorption coefficient,air permeability, air filtration efficiency, and thermal conductivity of the CMF. Anotherarea of study which can be significant to increase its commercial viability is working onreducing the incubation time of mycelia. In other words, finding the optimum incubationconditions to obtain the same amount of mycelia in shorter time is crucial to reduceproduction costs.40Bibliography[1] M. N. Belgacem and A. Gandini, Monomers, polymers and composites from renew-able resources. Elsevier, 2011.[2] R. D. Straughan and J. A. Roberts, “Environmental segmentation alternatives: alook at green consumer behavior in the new millennium,” Journal of consumer mar-keting, vol. 16, no. 6, pp. 558–575, 1999.[3] F. La Mantia and M. 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