EFFECT OF SUBSTRATE CONCENTRATION AND SCALE UP ON THE POLYGALACTURONASE PRODUCTION

Öz

Pectinases have been used for many industrial applications since long time ago. The largest industrial application of these enzymes is in fruit juice and wine production for the extraction, filtration and clarification and for the maceration of fruits and vegetables. They work by enzymatic breaking down of the cell wall.In this study it was aimed to use the previously optimized shake flask media formulation in batch mode 1 L scale serial bioreactor system and 5 L scale in order to investigate the effects of substrate concentration and scale on PG activity and biomass production. In conclusion it was observed that average PG activity (101.29 U/ml) obtained in 5L scale bioreactor experiments was higher than the maximum PG activity (88.55 U/ml) at 40 g/L orange peel (OP) concentration obtained in the 1 L scale substrate concentration experiment. Furthermore,PG activity increased with an increase in substrate concentration except for 60 g/L orange peel concentration. 

Anahtar Kelimeler

• scale-up,polygalacturonase,Aspergillus sojae,bioreactor,orange peel

SUBSTRAT KONSANTRASYONU VE ÖLÇEK BÜYÜTMENİN POLİGALAKTURONAZ ÜRETİMİNE ETKİSİ

Öz

Pektinazlar, uzun zaman önceden beri bir çok endüstriyel uygulama için kullanılmıştır. Bu enzimlerin en büyük endüstriyel uygulaması meyve suyu ve şarap üretiminde ekstraksiyon, berraklaştırma ve filtrasyon ayrıca meyve ve sebzelerin maserayonudur. Hücre duvarının enzimatik parçalanması yoluyla etki ederler. Bu çalışmada, substrat konsantrasyonunun ve ölçek büyütmenin PG aktivitesi ve biyokütle üretimi üzerindeki etkilerini araştırmak için daha önce erlenlerde optimize edilmiş medya formülasyonunun kesikli tip 1 L ölçekli seri biyoreaktör sistemi ve 5 L ölçekte kullanılması amaçlanmıştır. Sonuç olarak, 5 L ölçekli biyoreaktör deneylerinde elde edilen ortalama PG aktivitesinin (101,29 U/ml), 1 L ölçekli substrat konsantrasyonu deneyinden elde edilen 40 g/L portakal kabuğu konsantrasyonundaki maksimum PG aktivitesinden (88,55 U/ml) daha yüksek olduğu belirlenmiştir. Ayrıca 60 g/L portakal kabuğu konsantrasyonu hariç substrat konsantrasyonu arttıkçaPG aktivitesi de artmıştır.

Anahtar Kelimeler

• ölçek büyütme,poligalakturonaz,Aspergillus sojae,biyoreaktör,portakal kabuğu

Kaynakça

1. 1. Jayani, R.S., Saxena, S., Gupta R. (2005). Microbial pectinolytic enzymes: A review. Process Biochem, 40(9):2931-2944, doi: 10.1016/j.procbio.2005.03.026.
2. 2. Gummadi, S.N., Manoj, N., Kumar, D.S. (2007). Structural and Biochemical Properties of Pectinases. In Industrial Enzymes, Polaina, J., MacCabe, A.P., Eds.; Springer: Heidelberg, pp. 99-115.
3. 3. Malvessi, M., Ve Da Silveira, M.M. (2004). Influence of medium composition and pH on the production of polygalacturonases by Aspergillus oryzae. Braz Arch Biol Technol, 47(5):693-702.
4. 4. Nakkeeran, E., Gowthaman, M.K., Umesh-Kumar, S., Subramanian, R. (2012). Techno-economic Analysis of Processes for Aspergillus carbonarius Polygalacturonase Production. J Biosci Bioeng, 113(5):634–640. 5. Biz, A., Finkler, A.T.J., Pitol, L.O., Medina, B.S., Nadia K., Mitchell, D.A. (2016). Production of pectinases by solid-state fermentation of a mixture of citrus waste and sugarcane bagasse in a pilot-scale packed-bed bioreactor. Biochem Eng J, 111:54–62.
5. 6. Pandey, A., Soccol, C.R., Mitchell, D. (2000). New developments in solid state fermentation: I-bioprocesses and products. Process Biochem, 35(10):1153-1169, http://dx.doi.org/10.1016/S0032-9592(00)00152-7.
6. 7. Songulashvili, G., Spindler, D., Jimene´z-Tobo´n, G.A., Jaspers, C., Kerns, G., Penninckx, M.J. (2015). Production of a high level of laccase by submerged fermentation at 120-L scale of Cerrena unicolor C-139 grown on wheat bran. C. R. Biologies, 338:121–125, http://dx.doi.org/10.1016/j.crvi.2014.12.001.
7. 8. Oncu, S., Tari, C., Unluturk, S. (2007). Effect of various process parameters on morphology, rheology, and polygalacturonase production by Aspergillus sojae in a batch bioreactor. Biotechnol Prog, 23:836-845.
8. 9. Waites, M.J., Morgan, N.L., Rockey J.S., Gary H. (2001). Industrial Microbiology: An Introduction, Blackwell Science Ltd, pp. 107-108.

9. 10. Göğüş, N., Hakgüder Taze, B., Demir, H., Tarı, C., Ünlütürk, S., Lahore, M.F. (2014). Evaluation of orange peel, an industrial waste, for the production of Aspergillus sojae polygalacturonase considering both morphology and rheology effects. Turk J Biol, 38:537-548, doi:10.3906/biy-1308-47
10. 11. Panda, T., Naidu, G.S.N., Sinha, J. (1999). Multiresponse analysis of microbiological parameters affecting the production of pectolytic enzymes by Aspergillus niger: A statistical view. Process Biochem, 35, 187-195.
11. 12. Dubois, M., Gilles, K.A., Hamilton, J.K., Rebers, P.A., Smith, F. (1956). Colorimetric Method for Determination of Sugars and Related Substances. Anal Chem, 28(3):350-356.
12. 13. Bradford, M. (1976). A Rapid and Sensitive Method for the Quantitation of microgram Quantities of Protein Utilizing the Principle of Protein-dye Binding. Anal Biochem, 72:248-254.
13. 14. Maldonado, M.C., Strasser de Saad A.M. (1998). Production of pectinesterase and polygalacturonase by Aspergillus niger in submerged and solid state systems. J Ind Microbiol Biotechnol, 20(1):34–38, doi: 10.1038/sj.jim.2900470.
14. 15. Pollard, D.J., Kirschner, T.F., Hunt, G.R., Tong, I-T., Stieber, R., Salmon, P.M. (2006). Scale Up of a Viscous Fungal Fermentation: Application of Scale-Up Criteria with Regime Analysis and Operating Boundary Conditions. Biotechnol Bioeng, 96(2):307-317, doi:10.1002/bit.21112.
15. 16. Rocha-Valadez, J.A., Estrada, M., Galindo, E., Serrano-Carreon, L. (2006). From shake flasks to stirred fermentors: Scale-up of an Extractive Fermentation Process for 6-pentyl-a-pyrone Production by Trichoderma harzianum using Volumetric Power Input. Process Biochem, 41(6):1347–1352, doi:10.1016/j.procbio.2006.01.013.
16. 17. Junker, B.H., Hesse, M., Burgess, B., Masurekar, P., Connors, N., Seeley, A. (2004). Early Phase Process Scale-Up Challenges for Fungal and Filamentous Bacterial Cultures. Appl Biochem Biotechnol, 119(3):241-277, doi: 10.1007/s12010-004-0005-x.
17. 18. Junker, B., Walker, A., Hesse, M., Lester, M., Vesey, D., Christensen, J., Burgess, B., Connors, N. (2009). Pilot-scale process development and scale up for antifungal production Bioprocess Biosyst Eng, 32(4):443–458, doi: 10.1007/s00449-008-0264-y.
18. 19. Fontana, R.C., Polidoro, T.A., da Silveira, M.M. (2009). Comparison of stirred tank and airlift bioreactors in the production of polygalacturonases by Aspergillus oryzae. Bioresour Technol, 100(19):4493–4498, http://dx.doi.org/10.1016/j.biortech.2008.11.062.
19. 20. Meneghel, L., Reis, Guilherme, P., Reginatto, C., Malvessi, E., da Silveira, M.M. (2014). Assessment of pectinase production by Aspergillus oryzae in growth-limiting liquid medium under limited and non-limited oxygen supply. Process Biochem, 49:1800–1807, http://dx.doi.org/10.1016/j.procbio.2014.07.021.
20. 21. Wolf-Márquez, V.E., Martínez-Trujillo, M.A., Osorio, G.A., Patiño, F., Álvarez, M.S., Rodríguez, A., Sanromán, M.Á., Deive, F.J. (2017). Scaling-up and ionic liquid-based extraction of pectinases from Aspergillus flavipes cultures. Bioresour Technol, 225:326–335, http://dx.doi.org/10.1016/j.biortech.2016.11.067.