Malaysian Journal of Analytical Sciences Vol 21 No 1 (2017): 231 - 239

DOI: http://dx.doi.org/10.17576/mjas-2017-2101-27

 

 

 

SEED CULTURE CONDITIONS FOR HIGH YIELD OF CELLULOLYTIC ENZYMES PRODUCTION FROM Pycnoporus sanguineus

 

(Keadaan Kultur Benih untuk Penghasilan Enzim Selulolitik daripada Pycnoporus sanguineus)

 

Rafidah Jalil¹, Mohd Sahaid Kalil¹*, Norliza Abdul Rahman¹, Aidil Abdul Hamid², Mohd Farid Ahmad³

 

¹Department of Chemical & Process Engineering, Faculty of Engineering & Built Environment

²School of Bioscience and Biotechnology, Faculty of Science & Technology

Universiti Kebangsaan Malaysia, 43600 UKM Bangi, Selangor, Malaysia

³Forest Research Institute Malaysia (FRIM), 52109 Kepong, Selangor, Malaysia

 

*Corresponding author: sahaid@ukm.edu.my

 

 

Received: 21 October 2015; Accepted: 14 June 2016

 

 

Abstract

Low yields and productivity in fermentation industry usually due to poor quality of seed culture (inoculum). As the first stage in the fermentation process, inoculum consistency in terms of size and quality is clearly important for high yield of enzymes production. Selection of microorganism also plays an important role in improving the productivity. Fungi type of white rot basidiomycetes species is a well-known cellulolytic enzymes producer and capable to degrade many types of lignocellulosic biomass. Cellulolytic enzymes produced from white rot fungi, Pycnoporus sanguineus (PS) were investigated. The PS cultured on different agar media and parameter conditions of seed culture preparation in liquid medium broth were compared. Seed culture conditions of PS were influenced by many factors such as type of medium used for fungal growth, temperature, incubation time in liquid medium and subculture time on agar medium. PS was full-grown on potato dextrose agar (PDA) on day 5 compared to malt extract agar (MEA) on day 7. Seed culture conditions was determined using standard liquid medium, potato dextrose broth (PDB) at constant temperature (30 ^°C), agitation speed (150 rpm) and pH (4.8) for duration of seven days using two different subculture of PS that were grown for five and seven days. It was found that the five days’ subculture of PS shown 3.93% higher cell dry weight of fungal biomass (3.44 g/L on day 5) and higher cellulolytic enzymes activity; 95.49% (FPase), 3.14% (CMCase), 7.71% (exoglucanases) and 14.93% (xylanases). Maximum cellulolytic enzymes were found after 48 hours’ incubation with filter paper activity (FPase) of 1.79 U/mL, carboxymethyl cellulase activity (CMCase) of 3.36 U/mL, exoglucanase activity (Avicelase) of 0.59 U/mL and xylanase enzymes activity of 0.66 U/mL. Reducing sugars concentration decreased from day 1 to 7 due to consumption of sugars for fungal growth. Seed culture conditions were strongly influenced by subculture and incubation time in liquid medium to produce high yield of cellulolytic enzymes from PS.

 

Keywords:  white rot fungi, Pycnoporus sanguineus, seed culture, cellulolytic enzymes, fermentation

 

Abstrak

Hasil dan produktiviti yang rendah dalam industri fermentasi biasanya disebabkan oleh kultur benih (inokulum) yang kurang berkualiti. Pada peringkat pertama proses fermentasi, konsistensi inokulum dari segi saiz dan kualiti sangat penting bagi pengeluaran enzim yang tinggi. Pemilihan mikroorganisma juga memainkan peranan penting dalam meningkatkan produktiviti. Jenis kulat pereput putih dari spesies basidiomycetes sangat terkenal sebagai pengeluar enzim selulolitik dan ia mampu untuk mengurai pelbagai jenis bahan lignoselulosa. Dalam kajian ini, enzim selulolitik dihasilkan daripada kulat pereput putih, Pycnoporus sanguineus (PS) yang dihidupkan di atas agar dan perbandingan parameter berbeza penyediaan kultur benih dalam medium cecair dikaji. Kultur benih PS dipengaruhi oleh pelbagai faktor seperti jenis medium yang digunakan untuk pertumbuhan kulat, suhu, masa pengeraman di atas medium agar dan di dalam medium cecair. Pertumbuhan maksimum PS di atas agar kentang dekstrosa (PDA) dicapai pada hari ke-lima berbanding dengan agar ekstrak malt (MEA) pada hari ke-tujuh. Kultur benih ditentukan dengan menggunakan medium piawai iaitu medium kentang dekstrosa (PDB) pada suhu malar (30 ^°C), kelajuan pengadukan (150 rpm) dan pH (4.8) untuk tempoh masa tujuh hari menggunakan dua kultur PS berbeza yang dieram selama lima dan tujuh hari. Didapati bahawa kultur PS 5 hari menunjukkan berat kering sel kulat yang lebih tinggi iaitu 3.93% (3.44 g/L pada hari ke-lima) dan aktiviti enzim selulolitik lebih tinggi; 95.49% (FPase), 3.14% (CMCase), 7.71% (eksoglukanase) and 14.93% (xilanase). Pengeluaran enzim selulolitik maksimum diperolehi selepas 48 jam pengeraman dengan aktiviti kertas turas (FPase) 1.79 U/mL, aktiviti karboksimetil selulase (CMCase) 3.36 U/mL, aktiviti eksoglukanase (Avicelase) 0.59 U/mL dan aktiviti enzim xilanase 0.66 U/mL. Selain itu, kepekatan gula penurun berkurang dari hari 1-7 disebabkan oleh penggunaan gula untuk pertumbuhan kulat. Ini menunjukkan bahawa keadaan inokulum dipengaruhi oleh jenis kultur dan masa pengeraman dalam medium cecair untuk pengeluaran hasil enzim selulolitik yang tinggi dari PS.

 

Kata kunci:  kulat pereput putih, Pycnoporus sanguineus, kultur benih, enzim selulolitik, penapaian

 

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