DETERMINATION OF TANNASE ACTIVITY OF THE LACTIC ACID BACTERIA THAT ISOLATED FROM FERMENTED PRODUCTS

Abstract

In this study, 10 isolates obtained from fermented products in previous studies were identified byDuPont Qualicon RiboPrinter®system. Tannase activity was determined by spectrophotometric andvisual reading methods. Lactobacillus curvatus P3X showed the highest tannase activity. Approximatemolecular weight of the enzyme was determined by SDS- PAGE method. The effect of differenttemperatures, pH values, substrate concentrations, and the concentrations of tannic acid and variousminerals, on activity of tannase of L. curvatus P3X, of which showed high tannase activity, wereinvestigated. As a result of tests, the optimum temperature was found as 37°C, optimum pH was foundas 5.0, methyl gallate concentration that was used as the substrate was found as 7 mM and tannic acidconcentration was found as 1.75 mM. Tannase activity of lactic acid bacteria decreased in the presenceof Hg+2and Mg+2ions, Ca+2and Zn+2ions did not show any effect on tannase activity. In addition, it wasdetected that surfactant (Tween 80), chelate (EDTA), inhibitor (DMSO), and denaturing agents (urea)had no effect on tannase activity

Keywords

• Tannase, lactic acid bacteria, methyl gallate

FERMENTE GIDALARDAN İZOLE EDİLEN LAKTİK ASİT BAKTERİLERİNİN TANNAZ AKTİVİTESİNİN BELİRLENMESİ

Abstract

Bu çalışmada, daha önceki çalışmalarda fermente ürünlerden izole edilen 10 izolatın DuPont QualiconRiboPrinter®sistem ile tanımlamaları yapılmıştır. Spektrofotometrik ve görsel okuma metodu ile tannazaktivitesi belirlenmiştir. En yüksek aktiviteyi Lactobacillus curvatus P3X izolatı göstermiştir. L. curvatusP3X’e ait enzimin yaklaşık molekül ağırlığı SDS-PAGE yöntemi ile belirlenmiştir. Yüksek tannaz aktivitesigösteren L. curvatus P3X’in tannaz aktivitesi üzerine farklı sıcaklık, pH, substrat yoğunluğu, tannik asitkonsantrasyonu ve çeşitli minerallerin etkisi araştırılmıştır. Yapılan testler sonucunda optimum sıcaklık37°C, pH 5.0 substrat olarak kullanılan metil gallat konsantrasyonu 7 mM, tannik asit konsantrasyonuise 1.75 mM olarak bulunmuştur. Laktik asit bakterilerinin tannaz aktivitesi Hg+2ve Mg+2iyonlarınınvarlığında düşerken, Ca+2ve Zn+2’nun tannaz aktivitesi üzerine etki yapmadığı görülmüştür. Ayrıcayüzey aktif madde (Tween 80), şelat (EDTA), inhibitör (DMSO) ve yapıyı bozan maddenin de (Üre)tannaz aktivitesine tesir etmediği saptanmıştır

Keywords

• Tannaz, laktik asit bakterisi, metil galat

References

1. Rodriguez H, De las Rivas B, Gomez-Cordoves C, Munoz R. 2008. Degradation of Tannic Acid by Cell-Free Extracts of Lactobacillus plantarum. Food Chem, 107: 664-670.
2. Osawa R, Kuroiso K, Goto S, Shimizu A. 2000. Isolation of Tannin- Degrading Lactobacilli From Humans and Fermented Foods. Appl Environ Microbiol, 66 (7): 3093-3097.
3. Yao J, Guo GS, Ren GH, Liu YH. 2014. Production, Characterization and Applications of Tannase. J Mol Catal B: Enzym. 101: 137-147.
4. Temel O, Kıvanç M. 2015. Tannase Activity by Enterococcus faecium Isolated From Local Fermented Food. J Ferment Technol, 3:2.
5. Zhang S, Gao X, He L, Qiu Y, Zhu H, Cao Y. 2015. Novel Trends For Use of Microbial Tannases. Prep Biochem Biotech, 45: 221-232.
6. Kumar M, Beniwal V, Salar KR. 2015. Purification and Characterization of A Thermophilic Tannase From Klebsiella pneumoniae KP 715242. Biocatal Agric Biotech, 4: 745-751.
7. Vaquero I, Marcobal A, Munoz R. 2004. Tannase Activity by Lactic Acid Bacteria From Grape Must and Wine. Int J Food Microbiol, 96 (2): 199-204.
8. Nishitani Y, Osawa R. 2005. Deceptive Halo Formation by Tannase-Defective Bacteria on Tanin-Treated Plate Media. Microbes Environ, 20 (2): 117-119.

9. Nishitani Y, Osawa, R. 2003. A Novel Colorimetric Method to Quantify Tannase Activity of Viable Bacteria. J Microbiol Meth, 54: 281-284. 10. Kwon T, Shim S, Lee JH. 2008. Characterization of Lactobacilli With Tannase Activity Isolated From Kimchi. Food Sci Biotechnol, 17 (6): 1322-1326. 11. Rodriguez H, Rivas B, Cordoves CG, Munoz R. 2008. Characterization of Tannase Activity in Cell-Free Extracts of Lactobacillus plantarum CECT 748T. Int J Food Microbiol, 121: 92-98.
10. Madigan MT, Martinko MM, Stahl DA, Clark DP. (Ed). 2013. Brock Biology of Microorganisms, 13th Ed, Pearson Education Inc. San Francisco, CA 94111.USA.
11. Ayed A, Hamdi M. 2002. Culture Conditions of Tannase Production by Lactobacillus plantarum. Biotechnol Lett, 24: 1763-1765.
12. Kostinek M, Specht I, Edward VA, Pinto C, Egounlety M, Sossa C, Mbugua S, Dortu C, Thonart P, Taljaard L. 2007. Characterisation and Biochemical Properties of Predominant Lactic Acid Bacteria From Fermenting Cassava For Selection as Starter Cultures. J Food Microbiol, 114: 342-351.
13. Bhat TK, Singh B, Sharm OP. 1998. Microbial Degradation of Tannins; A Current Perspective. Biodegradation, 9: 343-357.
14. Mingshu L, Qiang H, Dongying J. 2006. Biodegradaton of Gallotannins and Ellagitannins. J Basic Microbiol, 46 (1): 68-84.
15. Skene IK. and Brooker JD. 1995. Characterization of Tannin Acylhydrolase Activity in The Ruminal Bacterium Selenomonas ruminantium. Anaerobe, 1 (6): 321-327.
16. Nishitani Y, Sasaki E, Fujisaw T. and Osawa R. 2004. Genotypic Analyses Of Lactobacilli With A Range of Tannase Activities Isolated From Human Feces and Fermented Foods. Syst Appl Microbiol, 27: 109-117.
17. Kar B, Banerjee R, Bhattacharyya BC. 2003. Effect of Additives on The Behaviorual Properties of Tannin Acyl Hydrolase. Process Biochem, 38: 1285-1293.
18. Sabu A, Kiran GS, Pandey A. 2005. Purification and Characterization of Tannin Acyl Hydrolase From Aspergillus niger ATCC 16620. Food Technol Biotech. 43: 133-138.
19. Fitzpatrick DE, Bing B, Maggi DA, Fleming RC, O'Malley RM. 2002. Vasodilating Procyanidins Derived From Grape Seeds. Ann New York Acad Sci, 957: 78-89.