Biodegradation of Three Agricultural Wastes by a White-rot Fungus Pleurotus pulmonarius (Fries) Quetlet. C. O. Adenipekun a and O. J.

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1 Nture nd Science 2013;11(2) Biodegrdtion of Three Agriculturl Wstes y White-rot Fungus Pleurotus pulmonrius (Fries) Quetlet C. O. Adenipekun nd O. J. Dd Deprtment of Botny, University of Idn, Idn, Nigeri oyinpek@yhoo.com Astrct: Studies were crried out on the degrdtion of cotton wste, rice strw nd coco pod husks using Pleurotus pulmonrius in cultures incuted for 0-60 dys. The proximte composition, percentges of nitrogen, cron, potssium nd phosphorus, ph, lignin contents, orgnic mtter nd enzyme production were determined using the sustrtes degrded y P. pulmonrius. Crude protein incresed significntly throughout the incution period from 1.27% in the control to 12.63% in cotton wste, 6.65% to 14.82% in rice strw nd in coco pod husk from 7.04% to 13.82%. Crude fire decresed significntly in cotton wste nd coco husk from 5.88% to 5.31% nd from 39.88% to 34.95% respectively ut n increse ws oserved in rice strw from 18.42% in control to 28.08% fter 60dys of incution period. The nutrient contents, ph vlues, orgnic mtter showed significnt differences of (P 0.05) in the three sustrtes s the dys of fermenttion incresed. Lignin contents of treted cotton wste nd coco husk were degrded y Pleurotus pulmonrius ut n increse ws oserved in the rice strw. Cellulse ctivities decresed significntly in contrst to the lignse ctivities where consistent increse ws oserved on the degrded sustrtes s the incution period incresed. [Adenipekun CO, Dd OJ. Biodegrdtion of three Agriculturl Wstes y White-rot Fungus Pleurotus pulmonrius (Fries) Quetlet. Nt Sci 2013;11(2):19-25]. (ISSN: ). 4 Key words: Degrdtion; Cotton wste; Rice strw; Coco pod husks; Pleurotus pulmonrius 1. Introduction Pleurotus pulmonrius, commonly known s the Indin oyster, Phoenix mushroom, or the Lung oyster, is mushroom very similr to Pleurotus ostretus, the perl oyster, ut with few noticele differences; the cps of P. pulmonrius re pler nd smller thn in P. ostretus (Stmets, 2000). P. pulmonrius in nture grow in clusters on ded wood nd cn e grown throughout the yer. Owing to the tolernce of Pleurotus pulmonrius to high temperture, it hs een reported tht its speedy fruiting nd yield efficiencies ttrct mny cultivtors to the mushroom (Trudell nd Ammirti, 2009). The white-rot fungus P. pulmonrius, is wood decying sidiomycetes which is cple of degrding not only lignin ut lso vrile reclcitrnt environmentl pollutnts due to its ility to secrete lignolytic enzymes such s lignin peroxidse, mngnese peroxidse nd lccses which id the degrdtion process (Ogo et l., 2006). Wste, generted from the mechnicl processing of rw cotton prior to spinning, provides n idel sustrte for the growth of some edile mushrooms notly Volvriell volvce the Chinese or strw mushroom nd the oyster mushrooms (Pleurotus spp). Cotton wste culture widely vrile mounts of totl nitrogen from % for Pleurotus (Chng, 1999). It hs iologicl efficiency of 56-86% for Pleurotus. Rice strw consists predominntly of cell wlls, comprised of cellulose, hemicellulose, nd lignin. To rek down these components cellulse, hemicellulse nd ligninse re required (Schiere nd Irhim, 1989). Huge quntities of gro-industril iomss including out 900 million tons of rice strw re produced worldwide nnully, more thn 90% eing produced in Asi (Jhromi et l., 2011). Coco husk is considered vlule source of dietry fire nd is gining considerle interest in economiclly dvnced countries. The knowledge of the qulity of the protein frction in the husk is impertive in developing nutritionl formultions in new products (Mkkr et l., 1997). Mushroom hyphe secrete lrge mounts of extrcellulr enzymes which ring out the degrdtion of mcromolecules such s cellulose, hemicellulose, lignin nd protein in the sustrtes (Nrsi et l., 2006; Kuforiji nd Fsidi, 2008). Adenipekun nd Okunlde (2012) investigted the potentil of Pleurotus ostretus in degrdtion of mize stovers nd rttn wood. Recycling of these unfvorle mterils through mushroom culture cn increse griculturl efficiency nd enhnce degrdtion process of ligno-cellulose sources (Oodi et l., 2003). Therefore, this study ws undertken to determine the potentil ility of cotton wste, rice strw nd coco husks eing degrded y Pleurotus pulmonrius. 2 Methods 2.1 The fungus Pure cultures of Pleurotus pulmonrius ws otined from the Plnt Physiology Lortory, 19

2 Nture nd Science 2013;11(2) Deprtment of Botny, University of Idn, Nigeri. Fresh cultures were otined y repeted suculturing on Potto Dextrose Agr (PDA). 2.2 Degrdtion of Sustrtes y P. pulmonrius The sustrtes used for this study were cotton wste, rice strw nd coco pod husk. Cotton wste (Gosspypium hirsutum) ws otined from Textile Mills, Idn. Freshly hrvested rice strw ws collected from Interntionl Institute of Tropicl Agriculture (I.I.T.A) in Idn. It ws sun-dried for two weeks to prevent decomposition. Fresh coco husks were otined nd processed t Coco Reserch Institute of Nigeri (CRIN) in Idn. 2.3 Preprtion of sustrtes The method of Adenipekun nd Fsidi (2005) ws employed. Twenty-five grms (25g) of ech dry wste were weighed into ech sterile ottle, squeezed out with muslin cloth nd 75ml distilled wter were dded. The ottles were immeditely covered with luminum foil nd sterilized in the utoclve t C for 15 minutes. The sustrtes were prepred in three replictes. 2.4 Inocultion Ech of the ottles ws inoculted t the centre of the sustrte with two gr plugs (7mm in dimeter) of vigorously growing myceli disc nd covered immeditely. They were kept in clen drk cupord in the lortory t 30 0 C nd 100% reltive humidity. The controls were dried in the oven t C for 48hours to determine the initil dry weight of the sustrtes (Bhrgv nd Orskov, 1987). The experiment ws replicted three times nd the ottles were hrvested fter 30 nd 60 dys nd their dry weights determined. 2.5 Anlyticl methods ph determintion: The ph of ech sustrte ws mesured with glss electrode of n electronic ph meter y dding 100ml distilled wter to 1g sustrte in clen ottles. After 18 hrs t room temperture, the ph of the suspension ws determined (Zdrzil nd Brunnert, 1982). The experiment ws done in triplicte. Lignin determintion: One grm (1g) of smple ws weighed into 250ml conicl flsk, to which 20ml of freshly prepred 72% H 2 SO 4 t 20 0 C ws dded nd crefully mixed then llowed to stnd for 2 hours. It ws lter refluxed with 238mls of distilled wter for 4hours. The insolule lignin ws llowed to settle overnight nd filtered. The residue ws then trnsferred into crucile of known weight nd dried in the oven t 60 0 C to constnt weight in desicctor nd weighed. Percentge lignin ws otined using the formul: % Lignin =weight of insolule lignin X 100 weight of oven dried smple 1 Nutrient content nlysis: Crude fire (CF), Crude protein (CP) nd ph were determined s descried y Zdrzil nd Brunnert (1982).The method of Assocition of Officil Agriculturl Chemists (A.O.A.C. 2003) were used to determine percentge orgnic cron, orgnic mtter, percentge nitrogen, phosphorus, potssium nd enzyme ssy. Enzyme ssy: Cellulse ws ssyed (Berridge, 1955) using 3,5-dinitroslicyclic cid (DNSA) nd the mount of reducing sugr formed ws determined. Lignse ws lso ssyed y the method of Berridge (1955). 2.6 Experimentl design nd Sttisticl nlysis A rndomised complete lock design ws used showing white-rot fungus, three sustrtes nd three incution periods were used. The white rot fungus ws P pulmonrius, the three sustrtes were cotton wste, rice strw nd coco pod husk while the incution period were 0, 30 nd 60 dys. An ANOVA tle ws prepred for ech, following the Duncn s multiple rnge test nd lest men significnt difference ws crried out on lignindegrding ilities of white-rot fungus. Ech tretment ws replicted three times. 3 Results 3.1 Effects of P. pulmonrius on proximte composition (g/100gdm) of degrded sustrtes Tle 1 shows the proximte composition (g/100gdm) of cotton wste, rice strw nd coco pod husk incuted for 0-60 dys. The dry mtter decresed with increse in incution period in treted cotton wste from 5.46% in the control to 3.70%, it lso decresed in rice strw from 3.90% in control to 3.19% nd coco pod husk from 5.10% in control to 3.75%. The crude fire decresed significntly compred to untreted sustrtes in cotton wste nd coco pod husk ut n increse ws oserved in rice strw s the rte of incution incresed. Cotton wste treted with P. pulmonrius decresed from 5.88% in the control to 5.31% fter 60 dys; coco pod husk treted with the fungus lso decresed significntly from 39.88% to 34.95% ut n increse from 18.42% in control to 28.08% ws oserved in rice strw fter 60 dys. The crude protein contents of the fungl treted sustrtes incresed significntly throughout the incution period, from 1.27% in the control to 12.63% in cotton wste, lso in rice strw from 6.65% (control) to 14.82% nd in coco pod husk from 7.04% in control to 13.82%. 3.2 Effects of P. pulmonrius on nutrient contents of degrded sustrtes 20

3 Nture nd Science 2013;11(2) The percentges of nitrogen, cron, potssium nd phosphorus of the three griculturl sustrtes re shown in Tle 2. With incresing incution period, the percentge nitrogen of treted cotton wste incresed from 0.24% in the control to 2.48%, in rice strw from 1.19% to 2.56% ut in coco pod husk it decresed from 1.26% to 0.22%. Cron content of treted cotton wste decresed from 47.37% in the control to 44.5% nd in coco pod husk from 37.22% to 31.87% while in rice strw, significnt increse ws oserved from 27.03% in the control to 41.69%. The potssium content in treted cotton wste decresed from 3.22% in control to 1.21%, ut incresed significntly in rice strw from 1.63% to 2.49% in control to 60 dys respectively while tht of coco pod husk incresed from 1.2% to 4.73%. Percentge phosphorus incresed significntly in the treted sustrtes. Phosphorus incresed from % in the control to % fter 2months of incution in cotton wste, % to % in rice strw nd % to % in coco husk ut of the three sustrtes treted with P. pulmonrius, rice strw hd the highest recorded vlue. 3.3 Effects of P. pulmonrius on orgnic mtter nd ph on degrded sustrtes Tle 3 shows the ph vlues of the treted sustrtes s the incution period incresed. The ph of cotton wste decresed from 5.86 to 5.53 ut in rice strw, the ph incresed from 5.77 to 6.42 nd in coco pod husk from 6.54 to 6.76 fter 60 dys incution. The percentge orgnic mtter incresed with increse in incution period of rice strw from 68.04% to 72.04% while decrese ws oserved in cotton wste nd coco pod husk from 81.91% in the control to 76.76% nd from 64.36% in the control to 55.10% respectively. 3.4 Effect of P. pulmonrius on lignin content of degrded sustrtes The percentges of lignin content of the sustrtes re shown in Tle 4. With incresing incution period, the lignin content of treted cotton wste degrded y the fungus reduced from 1.90% in the control to 1.45%, for coco pod husk it lso reduced from 28.76% to 22.33% ut in rice strw there ws n increse from 13.59% to 22.33%. 3.5 Effects of P. pulmonrius on degrded sustrtes for enzyme production Tle 5 shows the enzyme ctivities of P. pulmonrius on degrded cotton wste, rice strw nd coco husk over n incution period of 0-60 dys. It ws oserved tht cellulose ctivity decresed significntly ll through the period of incution on the three sustrtes from 9.07% to 8.47%, 9.50% to 7.27% nd 3.78% to 2.9% in cotton wste, rice strw nd coco pod husk respectively s compred with their controls. On the other hnd, consistent increse ws oserved in the lignse ctivity of the treted sustrte. Cotton wste incresed from 2.40% in the control to 2.97% fter 2months, similrly, rice strw incresed significntly from 2.83% to 5.03% nd coco husk lso incresed from 7.4% to 9.30% s compred with their control. 4. Discussion One gol of iologicl delignifiction using lignin degrding fungi is for the effective digestiility of the sustrte crohydrte. This cn e chieved y rpid lignin degrdtion (Adenipekun nd Fsidi, 2005). In this study, Pleurotus pulmonrius degrded cotton wste, rice strw nd coco pod husk. There ws consistent decrese in the dry mtter of the treted sustrte compred with the untreted one etween 0-60 dys. This is in line with (Jonthn et l., 2010) who reported tht dry mtter reduced significntly from 88.74% in control to 86.80% in Lentinus sunudus nd 86.55% in Pleurotus tuerregium. There ws significnt (P 0.05) increse in the crude protein (CP) contents of the sustrtes treted with P. pulmonrius with n increse in fermenttion dys. Similr result ws oserved in rice strw degrded y P. pulmonrius from 4.50% recorded in the control to 4.60% t dy 10, 4.78% t dy 20 nd 9.36% fter forty dys fermenttion (Jonthn et l., 2012). Similr trend ws lso oserved y Akinfemi et l., (2010) who reported tht the crude protein contents of the fungl treted sustrtes (sorghum stover) incresed for Pleurotus ostretus nd P. pulmonrius. The increse in crude protein contents might e due to the secretion of certin proteinous extr cellulr enzymes into the wste during their rekdown nd susequent metolism of the products (Kdiri, 1999). The decrese in crude fire (CF) of Pleurotus pulmonrius on degrded cotton wste nd coco husk could e due to cellulose enzymes eing secreted y cellulolytic fungi. Isikhuemhen nd Nerud (1999) oserved tht white rot fungi produced extrcellulr lignin modifying enzymes, the est chrcterized of which were lccse, lignin peroxidse nd mngnese peroxidses. Sfri et l. (2005) lso reported crude fier loss in whet, rley nd rice strw incuted with P. chrysosporium. Belewu nd Belewu (2005) s well reported tht degrdtion of nn leves decresed in fire contents or frctions due to the production of vrious enzymes during the vegettive nd reproductive phses with lignocelluloses degrding properties. In the opinion of Ad-Allh (2007) the high CP contents nd decrese CF frctions of iodegrded sustrtes could e the result of the fungl 21

4 Nture nd Science 2013;11(2) consumption of prt of the produced fermentle sugr produced y the ction of hydrolyzing enzyme of the fungus on the sustrte which then stimulted the fungus to produce high enzyme ctivities. A significnt increse in nitrogen percentge ws oserved in cotton wste nd rice strw nd decrese ws oserved in coco husk incuted with Pleurotus pulmonrius. This is similr to the findings of Anykorh nd Oltunji (2001) who reported higher nitrogen content (5.67%) in CW thn in CP (4.44%). Biologicl tretment reduces the ph of the fermented smples (Jhromi et l., 2011). The present study showed tht in cotton wste there ws reduction in ph over n incution period of 2months s compred with the control ut n increse ws oserved in rice strw nd coco pod husk over the sme period. The chnge in ph vlue my e ssocited with the increse in mino nitrogen content nd the presence of metolic wste products within the sustrtes. Fungi re generlly known to crry out their metolic ctivities t cidic ph (Fsidi, 1996). The present results show tht the orgnic mtter content ws higher (P 0.05) in rice strw nd this corresponds to the findings of Oziel et l., 2008 for Pleurotus-treted whet strw. In nother study, P. ostretus nd P. florid cultivted on rley strw, resulted in higher orgnic mtter digestiility for treted strw (Montñez et l., 2004). Esclon et l., (2001) reported tht sh concentrtion incresed 60 dys fter rley strws were incuted with P. ostretus, possily due to higher orgnic mtter used y the fungus. Lignin content decresed consistently (P 0.05) on Pleurotus-treted cotton wste nd coco husk in the present work. Removl of lignin contents lso hs contriution in the digestiility of lignocellulosic sustrtes (Zdrzil nd Brunnert, 1982). Lignin, cellulose nd hemicellulose frctions form the ulk of CPH fire. During the fermenttion process, the weekly chnges oserved in the fire frctions indicted the degree of lignocellulose iodegrdtion s well s the enzyme ctivities exhiited y P. ostretus on CPH (Alemwor et l., 2009). Cellulse ctivities decresed significntly t (P 0.05) over n incresing incution period. This result grees with the findings of (Adenipekun nd Okunlde, 2012) where they noted tht the depletion in enzymes production s the incution period incresed might e due to metolic ctivities during the process of degrdtion where enzymes were eing used up to id the process. On the other hnd, significnt increse ws oserved in the lignse ctivities of the treted sustrtes over the sme incution period. This oservtion is similr to tht of Kuforiji nd Fsidi (2008) who reported tht higher ctivities of proteinse, cellulse, lipse nd ctlse were oserved in the fruit odies compred to the scleroti. These enzymes were found to ffect the shelf life, food nutrient nd flvor of the mushroom. TABLE 1: Effects of P. pulmonrius on proximte composition (g/100gdm) of degrded sustrtes Sustrtes Incution dy period (dys) Dry mtter Crude fire Crude protein Cotton wste Rice strw c 6.65 Coco pod husk Ech vlue is men of 3 replictes. Mens with different superscripts in ech column re significntly different t P 0.05 ccording to Duncn s multiple rnge test 22

5 Nture nd Science 2013;11(2) Tle 2: Effects of Pleurotus pulmonrius on nutrient contents of degrded sustrtes Sustrtes Incution Orgnic Nitrogen (%) Potssium Phosphorus Period cron (%) (%) (%) (dys) Cotton wste Rice strw Coco pod husk c Ech vlue is men of 3 replictes. Mens with different superscripts in ech column re significntly different t P 0.05 ccording to Duncn s multiple rnge test. Tle 3: Effects of P. pulmonrius on orgnic mtter nd ph of degrded sustrtes Sustrtes Incution period (dys) Orgnic mtter (%) ph Cotton wste Rice strw Coco pod husk Ech vlue is men of 3 replictes. Mens with different superscript in ech column re significntly different t P 0.05 ccording to Duncn s multiple rnge test 23

6 Nture nd Science 2013;11(2) Tle 4: Effect of P. pulmonrius on lignin content of degrded sustrtes Sustrtes Incution period (dys) Lignin content Cotton wste Rice strw Coco pod husk Ech vlue is men of 3 replictes. Mens with different superscripts in ech column re significntly different t P 0.05 ccording to Duncn s multiple rnge test. Tle 5: Effects of P. pulmonrius on degrded sustrtes for enzymes production Sustrtes Incution period(dys) Lignse Unit/ml Cellulse Unit/ml Cotton wste Rice strw Coco pod husk Ech vlue is men of 3 replictes. Mens with different superscripts in ech column re significntly different t P 0.05 ccording to Duncn s multiple rnge test. References [1] Ad-Allh, S. A Biologicl tretments of some y products in ruminnts feeding. PhD Thesis. Al- Azhr University, Egypt 129pp. [2] Adenipekun, C. O. nd I. O. Fsidi, Degrdtion of selected griculturl wstes y Pleurotus tuerregium (Fries) Singer nd Lentinus sunudus (Berk). Nigerin edile mushroom. Advnces in Food Sciences, 27(2): [3] Adenipekun, C. O. nd Okunlde, O. A Biodegrdtion of Rttn Wood nd Mize Stovers y Pleurotus ostretus. Nture nd Science, 10 (5): [4] Akinfemi, A., Jonthn, S. G. nd Adu, O. A Improving the Nutritive vlue of millet stovers using iologicl tretment. Liyn Agriculturl Reserch Centre Journl Interntionl. 3: [5] Alemwor, F., Victori, P. D., Emmnuel, O. K. O. nd Jmes, H. O Effects of Pleurotus ostretus on coco pod husk composition: Influence of fermenttion period nd Mn 2+ supplementtion on the fermenttion process. Africn Journl of Biotechnology, 8(9):

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