Utilization of Cellulosic Bio Resource

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Utilization of cellulosicUtilization of cellulosic

bioresourcebioresource CelluloseCellulose -- a major source renewablea major source renewable

of carbon sourceof carbon source

Difficult to utilizeDifficult to utilize

Technology availableTechnology available economical?economical?



Major sources ofMajor sources of

cellulosicscellulosics Palm oil processingPalm oil processing

Rice productionRice production Timber processingTimber processing

Manucipal solid wastesManucipal solid wastes



Cellulosics from palm oilCellulosics from palm oil

processingprocessing



Cellulosics from palm oilCellulosics from palm oil

processingprocessing



Palm biomassPalm biomass carboncarbon

exploitationexploitation



Palm biomassPalm biomass aa

renewable resourcerenewable resource



Production of PHB fromProduction of PHB from

POMEPOME

Use lipolytic organismsUse lipolytic organisms produceproducelipases to breakdown lipids to fattylipases to breakdown lipids to fattyacids and glycerolacids and glycerol

Fatty acids then broken down throughFatty acids then broken down through F F--oxidation to produce acetyl CoAoxidation to produce acetyl CoA

Acetyl CoA produced incorporated into Acetyl CoA produced incorporated intoPHBPHB



Pathway for production of PHB Pathway for production of PHB using palm oil (Toh Mong Sahusing palm oil (Toh Mong Sah et alet al..

1995)1995)



Screening and isolation of PHA Screening and isolation of PHA

producers ( Amirulproducers ( Amirul et alet al. 2007). 2007)Soil samplesSoil samples

663 isolates663 isolates

Nile red stainingNile red staining

119 isolates were identified as possible PHA producers119 isolates were identified as possible PHA producers

cultivated in MSM containcultivated in MSM contain --butyrolactonebutyrolactone

95 isolates confirmed by GC analysis95 isolates confirmed by GC analysis

77 isolates produced homopolymer77 isolates produced homopolymer 18 isolates produced copolymer18 isolates produced copolymer

P(3HB)P(3HB) P(3HBP(3HB--coco--4HB)4HB)

USMAA1020USMAA1020



Observation PHA producersObservation PHA producers

under UV light microscopeunder UV light microscope

Individual cells when observed under UV light microscope. PHA

granules in the cell cytoplasm emitted orange fluorescence when

exposed to UV light.



Characterization of isolate USMAA1020Characterization of isolate USMAA1020

Morphological and biochemical

test resulted ;

Gram-negative

Aerobic

Non-spore forming

Motile

Rod-shaped

(0.7 ± 0.9 µm x 1.53 ± 3.0 µm)

Assimilates p-hydroxy-benzoate,

mesaconate, trans-aconitate,D/L-tartrate, citrate, malate,

adipate, caprate and gluconate

Growth at 30ºC and

37ºC but not at 42ºC

DNA G + C content is 62.3%

These results suggested that USM AA1020 belongs to the genus

Cupriavidus (Vandamme and Coenye, 2004).



Morphology of the isolateMorphology of the isolate

USMAA1020USMAA1020

Transmission electron micrograph of USM AA1020.

Morphology of cell with no PHA accumulation



TEM of USM AA 1020 showing morphology of cell with PHA granules in

the cell cytoplasm. PHA granules appear as electron-dense inclusions.

Morphology of the isolate USMAA1020Morphology of the isolate USMAA1020



Strategy for utilization ofStrategy for utilization of

cellulosicscellulosics Use enzyme cellulase for breakdownUse enzyme cellulase for breakdown

to produce glucoseto produce glucose

CellulaseCellulase a complex of 3 enzymesa complex of 3 enzymes

cellobiohydrolasecellobiohydrolase

endoglucanaseendoglucanase

F F--glucosidaseglucosidase



Sugar production fromSugar production from

cellulosicscellulosics Enzymes very expensiveEnzymes very expensive

PrePre--treatment of cellulosics required totreatment of cellulosics required tofacilitate hydrolysisfacilitate hydrolysis

Sugars produced can be utilized forSugars produced can be utilized forfermentation of various productsfermentation of various products



Mushroom productionMushroom production

A viable way to utilize cellulosics A viable way to utilize cellulosics

Has been shown to be commerciallyHas been shown to be commerciallyviableviable

Spent substrate can be used asSpent substrate can be used asfertilizerfertilizer



What are mushrooms?What are mushrooms?

Mushroom is a simple form of lifeMushroom is a simple form of lifeknown as fungusknown as fungus

Mushrooms are the edible fleshyMushrooms are the edible fleshyfruiting bodies of certain fungifruiting bodies of certain fungi

Mushrooms are popular for theirMushrooms are popular for their

delicacy and flavoured food valuedelicacy and flavoured food value Gathered wild or grown underGathered wild or grown under

cultivationcultivation



Varieties of mushrooms Varieties of mushrooms

commercially cultivatedcommercially cultivated Oyster mushroom (Oyster mushroom (PleurotusPleurotus sp.)sp.)

Paddy straw mushroomPaddy straw mushroom (Volvariella(Volvariellaspp.)spp.)

Shiitake (Shiitake (Lentinus edodesLentinus edodes))

Button mushroom (Button mushroom (Agaricus bisporus Agaricus bisporus))

Maitake (Maitake (Gifola frondosaGifola frondosa))



Steps in mushroomSteps in mushroom

cultivationcultivation Choosing a growing medium ie. substrate

Pasteurizing or sterilizing the medium

Seeding the beds with spawn

Maintaining optimal temperature, moisture,and other conditions for mycelium growth

and the conditions that favor fruiting - most challenging step

Harvesting, packaging, and marketing



Oyster mushroomOyster mushroom

cultivation flowcultivation flow--chartchartRaw material Chopping (3-5 cm) Soak straw (6-18 h)

Drain off excesswater

Fill-up gunnybags

Dip in hot water80-85oC x 15 min

Drain off excess

water

When cool to roomtemperature, fill bags in 3

layers of 10 cm each andsimultaneously spawn @2% inside sterile chamber



Oyster mushroomOyster mushroom

cultivation flowcultivation flow--chartchart Arrange bags in shelves(Spawn running requires20-25 days)

Pin head stage (takes 3-5days)

Cropping, 1st flush (3-4 daysduration after pin head ) andsubsequent flushes about 10days apart

Plucking on maturity(before spore shed starts)



What are potentialWhat are potential

substrates?substrates? Rice strawRice straw

Palm fibres?Palm fibres?

Saw dust?Saw dust?

Wood chips?Wood chips?



ReferencesReferences

MIAMIA Majid,Majid, DHDH Akmal, Akmal, LLLL FewFew.. A A Agustien, Agustien, MSMS Toh,Toh, MRMRSamian,Samian, NN NajimudinNajimudin andand MNMN Azizan Azizan ((19991999)).. ProductionProduction of of poly(poly(--33--hydroxybutyrate)hydroxybutyrate) byby ErwiniaErwinia spsp.. USMIUSMI--2020.. Int Int..

JournalJournal of of BiologicalBiological MacromoleculesMacromolecules 2525:: 9595--104104.. TohToh MongMong Sah,Sah, IsaIsa Abd Abd.. MajidMajid andand Azizan Azizan MohdMohd.. NoorNoor

((19951995)).. Aspects Aspects of of poly(poly(33--hydroxybutyrate)hydroxybutyrate) productionproduction bybyErwiniaErwinia spsp.. usingusing palmpalm oiloil asas thethe substratesubstrate.. 1818thth MalaysianMalaysianMicrobiologyMicrobiology SymposiumSymposium.. 2323--2424 August, August, Kuching,Kuching, SarawakSarawak..

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Utilization of Cellulosic Bio Resource