A New Approach to Prevent Enzymatic Browning in Sugar Beet: Herbiside Inhibitors

Yıl 2025, Cilt: 15 Sayı: 2, 624 - 633, 01.06.2025

Cansu Öztürk

https://doi.org/10.21597/jist.1568113

Öz

Enzymatic browning represents a significant challenge in the processing and storage of fresh produce within the food industry, as it can adversely affect consumer preferences. Various physical and chemical methods have been explored to inhibit the activity of polyphenol oxidase in order to extend the shelf life of products. In this study, it was aimed to prevent enzymatic browning in sugar beet (Beta vulgaris L. subsp. vulgaris) by limiting the polyphenol oxidase (PPO) activity with selected herbicides. Polyphenol oxidase enzyme was purified 46.35-fold by affinity chromatography using Sepharose-4B-L-tyrosine-p-aminobenzoic acid affinity gel. The molecular weight of the enzyme was found to be about 40 kDa by SDS-PAGE as a single band. Then, the inhibition effects of pendimethalin, bentazone, and glyphosate on purified PFO enzyme were investigated. IC50 values were determined as 0.157, 0.081, and 0.230 µM for pendimethalin, bentazone, and glyphosate herbicides, respectively. Kinetic assays indicated that bentazone was a remarkable inhibitor with the Ki value of 0.092 µM on PPO.

Anahtar Kelimeler

• Polyphenol oxidase , • purification , • inhibition , • herbicide

Şeker Pancarında Enzimatik Esmerleşmenin Önlenmesi İçin Yeni Bir Yaklaşım: Herbisit İnhibitörleri

Öz

Enzimatik esmerleşme, tüketici tercihlerini olumsuz etkileyebileceğinden, gıda endüstrisinde taze ürünlerin işlenmesi ve depolanmasında önemli bir zorluk teşkil etmektedir. Ürünlerin raf ömrünü uzatmak amacıyla, polifenoloksidaz enzim aktivitesini inhibe eden çeşitli fiziksel ve kimyasal yöntemler denenmiştir. Bu çalışmada, seçilmiş herbisitlerle polifenol oksidaz (PFO) aktivitesini sınırlayarak şeker pancarı (Beta vulgaris L. subsp. vulgaris) bitkisinde enzimatik esmerleşmenin önlenmesi amaçlanmıştır. Polifenol oksidaz enzimi, Sepharose-4B-L-tirozin-p-aminobenzoik asit afinite jeli kullanılarak afinite kromatografisi ile 46,35 kat saflaştırıldı. Enzimin molekül ağırlığı SDS-PAGE ile tek bant olarak yaklaşık 40 kDa olarak bulundu. Daha sonra, pendimetalin, bentazon ve glifosatın saflaştırılmış PFO enzimi üzerindeki inhibisyon etkileri araştırıldı. IC50 değerleri pendimetalin, bentazon ve glifosat herbisitleri için sırasıyla 0,157, 0,081 ve 0,230 µM olarak belirlendi. Kinetik analizler, bentazonun PFO üzerinde 0,092 µM'lik Ki değeriyle dikkate değer bir inhibitör olduğunu gösterdi.

Anahtar Kelimeler

Polifenol oksidaz , saflaştırma , inhibisyon , herbisit

Kaynakça

• Akdoğan, A., Divrikli, Ü., & Latif, E. L. Ç. İ. (2012). Pestisitlerin önemi ve ekosisteme etkileri. Akademik Gıda, 10(1), 125-132.
• Anh, T. M., Dzyadevych, S. V., Soldatkin, A. P., Chien, N. D., Jaffrezic-Renault, N., & Chovelon, J. M. (2002). Development of tyrosinase biosensor based on pH-sensitive field-effect transistors for phenols determination in water solutions. Talanta, 56(4), 627-634.
• Anh, T. M., Dzyadevych, S. V., Van, M. C., Renault, N. J., Duc, C. N., & Chovelon, J. M. (2004). Conductometric tyrosinase biosensor for the detection of diuron, atrazine and its main metabolites. Talanta, 63(2), 365-370.
• Arslan, O., Erzengin, M., Sinan, S., & Ozensoy, O. (2004). Purification of mulberry (Morus alba L.) polyphenol oxidase by affinity chromatography and investigation of its kinetic and electrophoretic properties. Food chemistry, 88(3), 479-484.
• Balcı, N., Türkan, F., Şakiroğlu, H., Aygün, A., & Şen, F. (2019). Purification and characterization of glutathione S-transferase from blueberry fruits (Vaccinium arctostaphylos L.) and investigated of some pesticide inhibition effects on enzyme activity. Heliyon, 5(4).
• Bradford, M. M. (1976). A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Analytical biochemistry, 72(1-2), 248-254.
• Cesko, C., Gashi, S., Arabaci, G., Palüzar, H., Durmishi, B., Bruci, E., ... & Özdemir, N. (2023). Investigation of the effects of pesticides on'Jonagold'apple (Malus x domestica) polyphenol oxidase enzyme activity. Turkish Journal of Agriculture and Forestry, 47(1), 57-66.
• Cheng, D., Wang, G., Tang, J., Yao, C., Li, P., Song, Q., & Wang, C. (2020). Inhibitory effect of chlorogenic acid on polyphenol oxidase and browning of fresh-cut potatoes. Postharvest Biology and Technology, 168, 111282. Çesko, C., Gashi, S., ARABACI, G., Palüzar, H., Durmishi, B., Bruci, E., ... & Özdemir, N. (2023). Investigation of the effects of pesticides on'Jonagold'apple (Malus x domestica) polyphenol oxidase enzyme activity. Turkish Journal of Agriculture and Forestry, 47(1), 57-66.
• Çiğdem, A., & Güller, U. (2022). Purification and characterization of catechol oxidase from Posof Badele apple (Malus domestica L): in vitro and in silico studies. International Journal of Food Engineering, 18(7), 513-524.
• Darko, G., & Akoto, O. (2008). Dietary intake of organophosphorus pesticide residues through vegetables from Kumasi, Ghana. Food and Chemical Toxicology, 46(12), 3703-3706.
• Fattouch, S., Raboudi-Fattouch, F., Ponce, J. V. G., Forment, J. V., Lukovic, D., Marzouki, N., & Vidal, D. R. (2010). Concentration dependent effects of commonly used pesticides on activation versus inhibition of the quince (Cydonia Oblonga) polyphenol oxidase. Food and chemical toxicology, 48(3), 957-963.
• Flurkey, W. H. (1986). Polyphenoloxidase in higher plants: immunological detection and analysis of in vitro translation products. Plant physiology, 81(2), 614-618.
• Fujita, S., Saari, N. B., Maegawa, M., Tetsuka, T., Hayashi, N., & Tono, T. (1995). Purification and properties of polyphenol oxidase from cabbage (Brassica oleracea L.). Journal of Agricultural and Food Chemistry, 43(5), 1138-1142.
• Gandía-Herrero, F., García-Carmona, F., & Escribano, J. (2004). Purification and characterization of a latent polyphenol oxidase from beet root (Beta vulgaris L.). Journal of Agricultural and Food Chemistry, 52(3), 609-615.
• Gerni, S., & Özdemir, H. (2024). Development of a new affinity chromatography method for purification of horseradish peroxidase enzyme. Biotechnology and Applied Biochemistry, 71(1), 202-212. Gonçalves, A. A., & de Oliveira, A. R. M. (2016). Melanosis in crustaceans: A review. LWT-Food Science and Technology, 65, 791-799.
• Jaśkiewicz, A., Kunicka-Styczyńska, A., Baryga, A., Gruska, R. M., Brzeziński, S., & Świącik, B. (2024). Evaluation of the Impact of an Enzymatic Preparation Catalyzing the Decomposition of Raffinose from Poor-Quality Beets during the White Sugar Production Process. Molecules, 29(15).
• Kaushik, G., Satya, S., & Naik, S. N. (2009). Food processing a tool to pesticide residue dissipation–A review. Food research international, 42(1), 26-40.
• Kaya, E. D. (2024). Biochemical properties of polyphenol oxidase purified from Sarali plum (Prunus domestica). Journal of Food Measurement and Characterization, 1-12.
• Kaya, E. D., & Bağci, O. (2021). Purification and biochemical characterization of polyphenol oxidase extracted from Kirmizi Kismis grape (Vitis vinifera L.). Journal of Food Biochemistry, 45(2), e13627.
• Koksal, Z., Kalin, R., Gulcin, I., & Ozdemir, H. (2018). Inhibitory effects of selected pesticides on peroxidases purified by affinity chromatography. International Journal of Food Properties, 21(1), 385-394.
• Laemmli, U. K. (1970). Cleavage of structural proteins during the assembly of the head of bacteriophage T4. nature, 227(5259), 680-685.
• Lazarević-Pašti, T., Momić, T., Radojević, M. M., & Vasić, V. (2013). Influence of organophosphorus pesticides on peroxidase and chlorination activity of human myeloperoxidase. Pesticide biochemistry and physiology, 107(1), 55-60.
• Li, F. (2020). Purification, kinetic parameters, and isoforms of polyphenol oxidase from “Xushu 22” sweet potato skin. Journal of Food Biochemistry, 44(11), e13452.
• Lin, H., Ng, A. W. R., & Wong, C. W. (2016). Partial purification and characterization of polyphenol oxidase from Chinese parsley (Coriandrum sativum). Food Science and Biotechnology, 25, 91-96.
• Lineweaver, H., & Burk, D. (1934). The determination of enzyme dissociation constants. Journal of the American chemical society, 56(3), 658-666.
• Mishra, B. B., and S. Gautam. "Polyphonel oxidases: biochemical and molecular characterization, distribution, role and its control. Enz Eng 5: 141." (2016).
• Moeini Alishah, M., Yıldız, S., Bilen, Ç., & Karakuş, E. (2023). Purification and characterization of avocado (Persea americana) polyphenol oxidase by affinity chromatography. Preparative Biochemistry & Biotechnology, 53(1), 40-53.
• Muñoz-Pina, S., Ros-Lis, J. V., Argüelles, Á., Coll, C., Martínez-Máñez, R., & Andrés, A. (2018). Full inhibition of enzymatic browning in the presence of thiol-functionalised silica nanomaterial. Food Chemistry, 241, 199-205.
• Nirmal, N. P., Benjakul, S., Ahmad, M., Arfat, Y. A., & Panichayupakaranant, P. (2015). Undesirable enzymatic browning in crustaceans: causative effects and its inhibition by phenolic compounds. Critical reviews in food science and nutrition, 55(14), 1992-2003.
• ÖZ, F., Colak, A., Özel, A., Sağlam Ertunga, Nagihan, N., & Sesli, E. (2013). Purification and characterization of a mushroom polyphenol oxidase and its activity in organic solvents. Journal of Food Biochemistry, 37(1), 36-44.
• Öztürk, C., & Küfrevioğlu, Ö. İ. (2024). Affinity gel synthesis from the p-aminobenzoic acid derivative 4-amino-2-methylbenzoic acid and purification of polyphenol oxidase from various plant sources. Protein Expression and Purification, 219, 106474.
• Öztürk, C., Aksoy, M., & Küfrevioğlu, Ö. İ. (2020). Purification of tea leaf (Camellia sinensis) polyphenol oxidase by using affinity chromatography and investigation of its kinetic properties. Journal of Food Measurement and Characterization, 14, 31-38.
• Öztürk, C., Bayrak, S., Demir, Y., Aksoy, M., Alım, Z., Özdemir, H., & İrfan Küfrevioglu, Ö. (2022). Some indazoles as alternative inhibitors for potato polyphenol oxidase. Biotechnology and Applied Biochemistry, 69(5), 2249-2256.
• Panadare, D.; Rathod, V.K. (2018). Extraction and purification of polyphenol oxidase: A review. Biocatal. Agric. Biotechnol, 14, 431–437.
• Raymond, J., Rakariyatham, N., & Azanza, J. L. (1993). Purification and some properties of polyphenoloxidase from sunflower seeds.
• Rung B, Schwack W. (2005). Aminoparathion: a highly reactive metabolite of parathion. 1. Reactions with polyphenols and polyphenol oxidase. Journal of Agricultural and Food Chemistry 53 (23): 9140–9145. https://doi.org/10.1021/ jf051520m
• Sae-leaw, T., & Benjakul, S. (2019). Prevention of melanosis in crustaceans by plant polyphenols: A review. Trends in Food Science & Technology, 85, 1-9.
• Shi, X., Tang, F., Zhou, X., & Bu, X. (2017). In vitro inhibition of polyphenol oxidase activity by insecticides and allelochemicals in Clostera anastomosis (Lepidoptera: Notodontidae) larvae and poplar trees. Journal of Entomological Science, 52(3), 239-247.
• Singh, A., & Wadhwa, N. (2017). Biochemical characterization and thermal inactivation of polyphenol oxidase from elephant foot yam (Amorphophallus paeoniifolius). Journal of Food Science and Technology, 54, 2085-2093.
• Taranto, F., Pasqualone, A., Mangini, G., Tripodi, P., Miazzi, M. M., Pavan, S., & Montemurro, C. (2017). Polyphenol oxidases in crops: Biochemical, physiological and genetic aspects. International journal of molecular sciences, 18(2), 377.
• Tinello, F., & Lante, A. (2018). Recent advances in controlling polyphenol oxidase activity of fruit and vegetable products. Innovative Food Science & Emerging Technologies, 50, 73-83.
• Yang, C. P., Fujita, S., Ashrafuzzaman, M. D., Nakamura, N., & Hayashi, N. (2000). Purification and characterization of polyphenol oxidase from banana (Musa sapientum L.) pulp. Journal of Agricultural and Food Chemistry, 48(7), 2732-2735.
• Yıldız, S., Bilen, Ç., & Karakuş, E. (2022). Purification of damson plum polyphenol oxidase by affinity chromatography and investigation of metal effects on enzyme activity. Preparative Biochemistry & Biotechnology, 52(9), 1019-1034.
• Zhang, S. (2023). Recent advances of polyphenol oxidases in plants. Molecules, 28(5), 2158.