ULTRASON DESTEKLİ ENZİMATİK YÖNTEMLE SARGASSUM VULGARE'DEN ANTİOKSİDAN PROTEİN EKSTRAKSIYONU: RSM İLE EKSTRAKSİYON PARAMETRELERİNİN OPTİMİZASYONU

Öz

Bu çalışmada, Sargassum vulgare'den proteinlerin ekstraksiyon koşulları optimize edilmiştir. Ekstraksiyon parametrelerinin (ultrases prop süresi: 0.09-2.91 dk ve enzim/substurat oranı (E/S): 0.18-1.02), protein miktarı (PM), toplam fenolik madde miktarı (TFMM) ve antioksidan aktivite (AOA) üzerine etkisini araştırmak ve optimizasyon çalışmalarını gerçekleştirmek için Box-Behnken Tasarım-Yanıt Yüzey Metodolojisi kullanılmıştır. Optimum protein ekstraksiyon koşulları, 2.5 dk ultrases prop uygulama süresi ve 0.90 E/S oranıdır. Optimum ekstraksiyon koşullarında, PM ve TFMM sırasıyla 248.30 mg protein/g kuru madde (km) ve 38.03 mg gallik asit eşdeğeri (GAE)/g km olarak bulunmuştur. Ayrıca AOA, CUPRAC yöntemi ile 53.77 mg Trolox eşdeğeri (TE)/g km ve ABTS yöntemi ile 19.88 mg TE/g km olarak belirlenmiştir. Bu bulgular, yüksek protein miktarı ve antioksidan aktivitesiye sahip makroalg protein ekstraktlarının gıda endüstrisi için potansiyelini araştıracak yeni çalışmalara bir temel oluşturabilir.

Anahtar Kelimeler

• Sargassum vulgare
• makroalg
• ekstraksiyon
• protein ekstraktı
• antioksidan aktivite

ULTRASOUND-ASSISTED ENZYMATIC EXTRACTION OF ANTIOXIDATIVE PROTEIN EXTRACTS FROM SARGASSUM VULGARE: OPTIMIZATION OF EXTRACTION PARAMETERS USING RSM

Öz

In this study, extraction conditions of proteins from Sargassum vulgare were optimized. The Box-Behnken design (BBD)-based Response Surface Methodology (RSM) was used to investigate and optimize the protein content (PC), total phenolic content (TPC), and antioxidant activity (AOA), which were affected by extraction parameters (ultrasonic probe time: 0.09-2.91 min and enzyme/substrate ratio (E/S): 0.18-1.02). The optimal extraction was achieved while applying an ultrasonic probe for 2.5 min and using an E/S of 0.90. Under this optimum conditions PC and TPC were found to be as 248.30 mg protein/g dry weight (dw) and 38.03 mg gallic acid equiavalent (GAE)/g dw, respectively. Moreover, AOA was determined to be 53.77 mg Trolox equivalent (TE)/g dw by CUPRAC and 19.88 mg TE/g dw by ABTS methods. These findings provide a good basis for future research into the potential of macroalgae protein extracts, which have a high protein content and antioxidant potential for food industry.

Anahtar Kelimeler

• Sargassum vulgare
• macroalgae
• extraction
• protein extract
• antioxidant activity.

Kaynakça

1. Acosta-Estrada, B. A., Gutiérrez-Uribe, J. A., Serna-Saldívar, S. O. (2014). Bound phenolics in foods, a review. Food Chemistry, 152, 46-55. https://doi.org/10.1016/j.foodchem.2013.11.093
2. Álvarez-Viñas, M., Rodríguez-Seoane, P., Flórez-Fernández, N., Torres, M. D., Díaz-Reinoso, B., Moure, A., Domínguez, H. (2021). Subcritical water for the extraction and hydrolysis of protein and other fractions in biorefineries from agro-food wastes and algae: A review. Food and Bioprocess Technology, 14, 373-387. https://doi.org/ 10.1007/s11947-020-02536-4
3. Apak, R., Güçlü, K., Özyürek, M., Karademir, S. E. N., Altun, M. (2005). Total antioxidant capacity assay of human serum using copper (II)-neocuproine as chromogenic oxidant: the CUPRAC method. Free Radical Research, 39(9), 949-961. https://doi.org/10.1080/ 10715760500210145
4. Arguelles, E. D. L. R., Monsalud, R. G., Sapin, A. B. (2019). Chemical composition and in vitro antioxidant and antibacterial activities of Sargassum vulgare C. Agardh from Lobo, Batangas, Philippines. International Society for Southeast Asian Agricultural Sciences, 25, 112-122.
5. Barclay, L. R. C., andVinqvist, M. R. (2003). Phenols as antioxidants. The Chemistry of Phenols, 839-908. https://doi.org/10.1002/ 0470857277.ch12
6. Bleakley, S., and Hayes, M. (2017). Algal proteins: extraction, application, and challenges concerning production. Foods, 6(5), 33. https://doi.org/ doi:10.3390/foods6050033
7. Bonilla Loaiza, A. M., Rodríguez-Jasso, R. M., Belmares, R., López-Badillo, C. M., Araújo, R. G., Aguilar, C. N., ... Ruiz, H. A. (2022). Fungal Proteins from Sargassum spp. using solid-state fermentation as a green bioprocess strategy. Molecules, 27(12), 3887.
8. Bozdemir, A., Şensu, E., Okudan, E. Ş., Özçelik, B., Yücetepe, A. (2022). Ultrasound-assisted enzymatic extraction of proteins from Gracilaria dura: Investigation of antioxidant activity and techno‐functional properties. Journal of Food Processing and Preservation, 46(8), e16803. https://doi.org/10.1111/jfpp.16803

9. Chan, P. T., and Matanjun, P. (2017). Chemical composition and physicochemical properties of tropical red seaweed, Gracilaria changii. Food Chemistry, 221, 302-310. https://doi.org/ 10.1016/j.foodchem.2016.10.066
10. de Melo, N. S. M., Cardoso, L. G., de Castro Nunes, J. M., Brito, G. B., Caires, T. A., de Souza, C. O., ... Druzian, J. I. (2021). Effects of dry and rainy seasons on the chemical composition of Ulva fasciata, Crassiphycus corneus, and Sargassum vulgare seaweeds in tropical environment. Brazilian Journal of Botany, 44(2), 331-344. https://doi.org/10.1007/s40415-021-00700-4
11. Ferruzzi, M. G., and Green, R. J. (2006). Analysis of catechins from milk–tea beverages by enzyme assisted extraction followed by high performance liquid chromatography. Food Chemistry, 99, 484–491
12. Field, L. M., Fagerberg, W. R., Gatto, K. K., Anne Böttger, S. (2017). A comparison of protein extraction methods optimizing high protein yields from marine algae and cyanobacteria. Journal of Applied Phycology, 29, 1271-1278. https://doi.org/ 10.1007/s10811-016-1027-9
13. Gressler, V., Yokoya, N. S., Fujii, M. T., Colepicolo, P., Mancini Filho, J., Torres, R. P., Pinto, E. (2010). Lipid, fatty acid, protein, amino acid and ash contents in four Brazilian red algae species. Food Chemistry, 120(2), 585-590. https://doi.org/10.1016/j.foodchem.2009.10.028
14. Harnedy, P. A., and FitzGerald, R. J. (2011). Bioactive proteins, peptides, and amino acids from macroalgae (1). Journal of Phycology, 47(2), 218–232. https://doi.org/10.1111/j.1529-8817.2011.00969.x
15. Joubert, Y., and Fleurence, J. (2008). Simultaneous extraction of proteins and DNA by an enzymatic treatment of the cell wall of Palmaria palmata (Rhodophyta). Journal of Applied Phycology, 20, 55-61. https://doi.org/10.1007/s10811-007-9180-9
16. Karkhane, M., Lashgarian, H. E., Mirzaei, S. Z., Ghaffarizadeh, A., Sepahvand, A., Marzban, A. (2020). Antifungal, antioxidant and photocatalytic activities of zinc nanoparticles synthesized by Sargassum vulgare extract. Biocatalysis and Agricultural Biotechnology, 29, 101791. https://doi.org/ 10.1016/j.bcab.2020.101791
17. Khaled, N., Hiba, M., Asma, C. (2012). Antioxidant and antifungal activities of Padina pavonica and Sargassum vulgare from the Lebanese Mediterranean Coast. Advances in Environmental Biology, 6(1), 42-48.
18. Khan, W., Rayirath, U. P., Subramanian, S., Jithesh, M. N., Rayorath, P., Hodges, D. M., ... Prithiviraj, B. (2009). Seaweed extracts as biostimulants of plant growth and development. Journal of Plant Growth Regulation, 28, 386-399. https://doi.org/10.1007/s00344-009-9103-x
19. Kumar, K. S., Ganesan, K., Selvaraj, K., Rao, P. S. (2014). Studies on the functional properties of protein concentrate of Kappaphycus alvarezii (Doty) Doty–An edible seaweed. Food Chemistry, 153, 353-360. https://doi.org/10.1016/ j.foodchem.2013.12.058
20. Liadakis, G. N., Tzia, C., Oreopoulou, V., Thomopoulos, C. D. (1995). Protein isolation from tomato seed meal, extraction optimization. Journal of Food Science, 60(3), 477-482. https://doi.org/10.1111/j.1365-2621.1995.tb09807.x
21. Lowry, O. H., Rosebrough, N. J., Farr, A. L., Randall, R. J. (1951). Protein measurement with the Folin phenol reagent. Journal of Biological Chemistry, 193, 265-275.
22. Mahmoud, S. H., Salama, D. M., El-Tanahy, A. M., Abd El-Samad, E. H. (2019). Utilization of seaweed (Sargassum vulgare) extract to enhance growth, yield and nutritional quality of red radish plants. Annals of Agricultural Sciences, 64(2), 167-175. https://doi.org/10.1016/j.aoas.2019.11.002
23. Mason, T. J., Paniwnyk, L., Lorimer, J. P. (1996). The uses of ultrasound in food technology. Ultrasonics Sonochemistry, 3(3), S253-S260. https://doi.org/10.1016/S1350-4177(96)00034-X
24. Miller, N. J., and Rice-Evans, C. A. (1997). Factors influencing the antioxidant activity determined by the ABTS•+ radical cation assay. Free Radical Research, 26(3), 195-199. https://doi.org/10.3109/10715769709097799
25. Moein, S., Moein, M., Ebrahimi, N., Farmani, F., Sohrabipour, J., Rabiei, R. (2015). Extraction and determination of protein content and antioxidant properties of ten algae from Persian Gulf. International Journal of Aquatic Science, 6(2), 29–38.
26. Moorthy, I. G., Maran, J. P., Muneeswari, S., Naganyashree, S., Shivamathi, C. S. (2015). Response surface optimization of ultrasound assisted extraction of pectin from pomegranate peel. International Journal of Biological Macromolecules, 72, 1323-1328. https://doi.org/10.1016/ j.ijbiomac.2014.10.037
27. Morais, H. A., Silvestre, M. P. C., Silva, M. R., Silva, V. D. M., Batista, M. A., Simões e Silva, A. C., Silveira, J. N. (2015). Enzymatic hydrolysis of whey protein concentrate: effect of enzyme type and enzyme: substrate ratio on peptide profile. Journal of Food Science and Technology, 52, 201-210. https://doi.org/10.1007/s13197-013-1005-z
28. Nursid, M., Khatulistiani, T. S., Noviendri, D., Hapsari, F., Hardiyati, T. (2020, May). Total phenolic content, antioxidant activity and tyrosinase inhibitor from marine red algae extract collected from Kupang, East Nusa Tenggara. In IOP Conference Series: Earth and Environmental Science (Vol. 493, No. 1, p. 012013). IOP Publishing.
29. Perumal, B., Chitra, R., Maruthupandian, A., Viji, M. (2019). Nutritional assessment and bioactive potential of Sargassum polycystum C. Agardh (Brown Seaweed). Indian Journal of Geo Marine Sciences, 48 (04), 492-498
30. Prasedya, E. S., Martyasari, N. W. R., Abidin, A. S., Ilhami, B. T. K., Padmi, H., Widyastuti, S., ... Sunarpi, H. (2021, March). Antioxidant activity of brown macroalgae Sargassum ethanol extract from Lombok coast, Indonesia. In IOP Conference Series: Earth and Environmental Science (Vol. 712, No. 1, p. 012038). IOP Publishing.
31. Ramakrishnan, V. V., Ghaly, A. E., Brooks, M. S., Budge, S. M. (2013). Extraction of proteins from mackerel fish processing waste using alcalase enzyme. Bioprocess Biotech, 3(2), 2-10.
32. Rodrigues, D., Freitas, A. C., Pereira, L., Rocha-Santos, T. A., Vasconcelos, M. W., Roriz, M., ... Duarte, A. C. (2015). Chemical composition of red, brown and green macroalgae from Buarcos bay in Central West Coast of Portugal. Food Chemistry, 183, 197-207. https://doi.org/ 10.1016/j.foodchem.2015.03.057
33. Sirbu, R., Teagu, N. P., Mirea, M., Negreanu-Pirjol, B. S. (2020). Bioactive compounds from three green algae species along Romanian Black sea coast with therapeutically properties. European Journal of Natural Sciences and Medicine, 3(1), 67-86. https://doi.org/10.26417/720mnv73y
34. Suwal, S., Perreault, V., Marciniak, A., Tamigneaux, É., Deslandes, É., Bazinet, L., ... Doyen, A. (2019). Effects of high hydrostatic pressure and polysaccharidases on the extraction of antioxidant compounds from red macroalgae, Palmaria palmata and Solieria chordalis. Journal of Food Engineering, 252, 53-59. https://doi.org/ 10.1016/j.jfoodeng.2019.02.014
35. Tekin, K., Akalın, M. K., Şeker, M. G. (2015). Ultrasound bath-assisted extraction of essential oils from clove using central composite design. Industrial Crops and Products, 77, 954-960. https://doi.org/10.1016/j.indcrop.2015.09.071
36. Taboada, C., Millán, R., Míguez, I. (2010). Composition, nutritional aspects and effect on serum parameters of marine algae Ulva rigida. Journal of the Science of Food and Agriculture, 90(3), 445-449. https://doi.org/10.1002/jsfa.3836
37. Thoo, Y. Y., Ho, S. K., Liang, J. Y., Ho, C. W., Tan, C. P. (2010). Effects of binary solvent extraction system, extraction time and extraction temperature on phenolic antioxidants and antioxidant capacity from mengkudu (Morinda citrifolia). Food Chemistry, 120(1), 290-295. https://doi.org/10.1016/j.foodchem.2009.09.064
38. Toor, R. K., and Savage, G. P. (2006). Changes in major antioxidant components of tomatoes during post-harvest storage. Food Chemistry, 99(4), 724–727. https://doi.org/10.1016/ j.foodchem.2005.08.049
39. Vásquez, V., Martínez, R., Bernal, C. (2019). Enzyme-assisted extraction of proteins from the seaweeds Macrocystis pyrifera and Chondracanthus chamissoi: Characterization of the extracts and their bioactive potential. Journal of Applied Phycology, 31(3), 1999-2010. https://doi.org/10.1007/ s10811-018-1712-y
40. Yuan, Y., Zhang, J., Fan, J., Clark, J., Shen, P., Li, Y., Zhang, C. (2018). Microwave assisted extraction of phenolic compounds from four economic brown macroalgae species and evaluation of their antioxidant activities and inhibitory effects on α-amylase, α-glucosidase, pancreatic lipase and tyrosinase. Food Research International, 113, 288-297. https://doi.org/ 10.1016/j.foodres.2018.07.021
41. Yucetepe, A., Okudan, E. Ş., Özçelik, B. (2022). Antioxidant activity and techno-functional properties of protein extracts from Caulerpa prolifera: Optimization of enzyme-assisted extraction by response surface methodology. Journal of Food & Nutrition Research, 61(3), 264-276.