بررسی فعالیت آنتیاکسیدانی محصولات هیدرولیز پروتئین بادام زمینی توسط آنزیمهای پپسین و آلکالاز
محورهای موضوعی : میکروبیولوژی مواد غذایی
هانیه حاجی کاظمی
1
(دانشجوی کارشناسی ارشد، گروه علوم و صنایع غذایی، واحد شهر قدس، دانشگاه آزاد اسلامی، تهران، ایران)
مهتا میرزایی
2
(استادیار گروه علوم و صنایع غذایی، واحد شهر قدس، دانشگاه آزاد اسلامی، تهران، ایران)
سعید میردامادی
3
(استاد پژوهشکده زیست فناوری، سازمان پژوهش های علمی و صنعتی ایران، تهران، ایران)
کلید واژه: آلکالاز, بادام زمینی, پپسین, فعالیت آنتی اکسیدانی, هیدرولیز آنزیمی,
چکیده مقاله :
مقدمه: پپتیدهای زیست فعال در ساختار پروتئین به صورت غیرفعال وجود داشته اما پس از پروتئولیز توانایی کاهش فشارخون، اثراتضدانعقادی، آنتیاکسیدانی، آرام بخشی، تاثیر بر سیستم ایمنی بدن و تاثیرات ضدمیکروبی از خود نشان میدهند. بادام زمینی یکی از مغزهایرایج مصرفی میباشد که میزان پروتئین بالایی داشته که میتواند به عنوان منبع استخراج پپتیدهای زیست فعال با توالی جدید و عملکردخاص مورد بررسی قرار گیرد.مواد و روشها: فعالیت آنتیاکسیدانی محصولات حاصل از هیدرولیز پروتئین بادام زمینی خام بوسیله آنزیمهای پپسین و آلکالاز درزمانهای مختلف هیدرولیز مورد بررسی قرار گرفت. پروتئین استخراج شده از بادام زمینی با روش استخراج با آب، به مدت 5 ساعت در معرضهیدرولیز بوسیله آنزیمهای پپسین و آلکالاز) با نسبت آنزیم به سوبسترا 1:10 ( به ترتیب در دمای 37 و 60 درجه سانتی گراد و pH برابر 2 و5/ 8 قرار گرفت سپس در طی زمان، پیشرفت هیدرولیز آنزیمی با روش OPA و فعالیت آنتیاکسیدانی با روشهای مهارکنندگی رادیکالهایDPPH و ABTS مورد بررسی قرار گرفتند.یافتهها: مقادیر گروههای آمین آزاد تولید شده توسط آنزیمهای پپسین و آلکالاز به ترتیب از مقادیر μM leu/mg protein 415 / 0 و 167 / 0 درزمان صفر به μM leu/mg protein517 / 0 و 263 / 0 بعد از 5 ساعت هیدرولیز رسیدند. حداکثر فعالیت مهارکنندگی رادیکال DPPH درمحصول هیدرولیز پپسین و آلکالاز به ترتیب mM TE/mg protein 2751 / 0 و 3644 / 0 و حداکثر فعالیت مهارکنندگی رادیکال ABTSبرای دو آنزیم mM TE/mg protein756 / 0 و 087 / 1 اندازه گیری شد.نتیجهگیری: نتایج نشان دادند که آنزیمهای پپسین و آلکالاز دارای توانایی هیدرولیز پروتئینهای بادامزمینی و تولید پپتیدهایآنتیاکسیدان هستند. نتایج حاصل از این تحقیق نشاندهنده پتانسیل استفاده از محصول هیدرولیز پروتئین بادام زمینی بوسیله آنزیمهای پپسینو آلکالاز، در فرمولاسیون غذاهای فراسودمند میباشد.
Introduction: The use of protein hydrolysate containing antioxidant peptides in the formulation of functional food has been increasing recently. The types and sequences of amino acids, the type of hydrolyzing enzymes and hydrolysis progress have some important impacts on the properties of protein hydrolysate. Materials and Methods: In this research, the effects of pepsin and alcalase enzymes (E/S:1/10) under optimal conditions of each ones, were investigated on the extraction of antioxidant peptides from peanuts protein. Peanut’s oil was extracted using solvent extraction method and protein was precipitated at isoelectric point. The extracted protein was subjected to the pepsin and alcalase enzymes for maximum period of five hours. The progress of hydrolysis was considered every thirty-minutes using Ortho-Phthalaldehyde (OPA) method. Results: The results indicated that the most hydrolysis occurs after 250 and 90 min of hydrolysis for pepsin and alcalase, respectively and the values of free amino acid groups increased from 167.0 to 263.0 μM leucin/mg protein (for alcalase) and from 415.0 to 517.0 μM leucin/mg protein (for pepsin). Moreover, the antioxidant activity of protein hydrolysate was investigated based on DPPH and ABTS radical scavenging activity. By increasing the degree of hydrolysis, the DPPH and ABTS radical scavenging activity increased simultaneously. The maximum values of DPPH and ABTS free radicals scavenging activity were measured respectively, 5175.0 and 756.0 mMTE/mg proteins (for pepsin) and 3644.0 and 1087.0 mMTE/mg proteins (for alcalase). Conclusion: The results indicated that the progress of enzymatic hydrolysis of peanut protein by alcalase and pepsin enzymes leads to producing more antioxidant peptides and the final products obtained can be considered as a candidate for producing functional foods.
Bamdad, F., Wu, J. & Chen, L. (2011). Effects of Enzymatic Hydrolysis on Molecular Structure and Antioxidant Activity of Barley Hordein. Journal of Cereal Science, 54, 20-28.
Bondet, V., Brand-Williams, W. & Berset, C. (1997). Kinetics and Mechanisms of Antioxidant Activity Using the DPPH. Free Radical Method. LWT-Food Science and Technology, 30(6), 609-615.
Chiang, W. D., Shih, C. J. & Chu, Y. H. (1999). Functional Properties of Soy Protein Hydrolysate Produced from a Continuous Membrane Reactor System. Food Chemistry, 65(2), 189-194.
Dryáková, A., Pihlanto, A., Marnila, P., Čurda, L. & Korhonen, H. J. (2010). Antioxidant Properties of Whey Protein Hydrolysates as Measured by Three Methods. European Food Research and Technology, 23 (6), 865- 874.
Fitznar, H. P., Lobbes, J. M. & Kattner, G. (1999). Determination of Enantiomeric Amino acids with High-Performance Liquid Chromatography and Pre-column Derivatisation with O-phthaldialdehyde and N-Isobutyrylcysteine in Seawater and Fossil Samples (mollusks). Journal of Chromatography, A832(1), 123-132.
He, R., Alashi, A., Malomo, S. A., Girgih, A. T., Cha D., Ju, X. & Aluko, R. E. (2013). Antihypertensive and Free Radical Scavenging Properties of Enzymatic Rapeseed Protein Hydrolysates. Food Chemistry, 141(1), 153-159.
Hou, R. Z. Y., Yang, G. Li, Y. B., Huang, H., Wang, Y. J., Liu, L. & Zhang, X. Z. (2006). Synthesis of a Precursor Dipeptide of RGDS (Arg‐Gly‐Asp‐Ser) Catalysed by the Industrial Protease Alcalase. Biotechnology and Applied Biochemistry, 44(2), 73-80.
Lowry, O., Rosebrough, N., Farr, A. & Randall, R. (1951). Protein Estimation by Lowry’s Method. The Journal of Biological Chemistry, 193(1), 265-275.
Ludescher, R. D. (1996). Physical and Chemical Properties of Amino Acids and Proteins. Food Proteins: Properties and Characterization: Wiley-VCH, PP. 23-70.
Makinen, S. (2014). Production, Isolation and Characterization of Bioactive Peptides with Antihypertensive Properties from Rapeseed and Potato Protein, phD thesis, University of turku.
Martysiak-Zurowska, D. & Wenta, W. (2012). A Comparison of ABTS and DPPH Methods for Assessing the Total Antioxidant Capacity of Human Milk. Acta Scientiarum Polonorum. Technologia Alimentaria, 11(1), 83-89.
Moure, A., Domínguez, H. & Parajó, J. C. (2006). Antioxidant Properties of Ultrafiltration-Recovered Soy Protein Fractions from Industrial Effluents and Their Hydrolysates. Process Biochemistry, 41(2), 447-456.
Morr, C. V., German, B., Kinsella J. E., Regenstein, J., Van Buren, J. P., Kilara, A., Lewis, B. A. & Mangino, M. E. (1985). A Collaborative Study to Develop a Standardized Food Protein Solubility Procedure. Journal of Food Science 50(6): 1715-1718.
Nwokolo, E. & Smartt, J. (1996). Food and Feed from Legumes and Oilseeds, Chapman & Hall London, UK.
Panyam, D. & Kilara, A. (1996). Enhancing the Functionality of Food Proteins by Enzymatic Modification. Trends in Food Science & Technology, 7(4), 120-125.
Quist, E. E. (2005). Peanut (Arachis hypogaea L.) as a Sourse of Antihypertensive and Antimicrobial Peptides. MSc thesis, University of Ghan.
Re, R., Pellegrini, N., Proteggente, A., Pannala, A., Yang, M. & .Rice-Evans, C. (1999). Antioxidant Activity Applying an Improved ABTS Radical Cation Decolorization Assay. Free Radical Biology and Medicine, 26(9), 1231-1237.
Saito, Y., Wanezaki, K., Kawato, A. & Imayasu, S. (1994). Structure and Activity of Angiotensin I Converting Enzyme Inhibitory Peptides from Sake and Sake Lees. Bioscience, Biotechnology, and Biochemistry, 58(10), 1767-1771.
Salunkhe,, D. K. (1992). World Oilseeds, Springer Science & Business Media.
Siow, H. L. & Gan, C. Y. (2013). Extraction of Antioxidative and Antihypertensive Bioactive Peptides from Parkia speciosa Seeds. Food Biochemistry, 141, 3435–3442.
Taheri, A., Jalalinezhad, S. & Anvar, S. A. A. (2012). Antihypertensive and Antioxidant Properties of Five Different Protein Hydrolysates Produced from Indian White Shrimp (Penaeus indicus) By-products. Journal of Comparative Pathobiology, 9 (1), 599-608 [In Persian].
Tang, C. H., Wang, X. S. & Yang, X. Q. (2009). Enzymatic Hydrolysis of Hemp (Cannabis sativa L.) Protein Isolate by Various Proteases and Antioxidant Properties of the Resulting Hydrolysates. Food Chemistry, 141, 1484–1490.
Turner, J. & Backman, P. (1991). Factors Relating to Peanut Yield Increases after Seed Treatment with Bacillus Subtilis. Plant Disease, 75(4): 347-353.
Wieser, H. (2007). Chemistry of Gluten Proteins. Food Microbiology, 24(2), 115-119.
Woodroof, J. (1983). Peanut Butter. Peanuts: Production, Processing and Products, AVI Publishing co, Inc, 3rd edn, West port, Connecticut 127.
You, L., Zhao, M., Cui, C., Zhao, H. & Yang, B. (2009). Effect of Degree of Hydrolysis on the Antioxidant Activity of Loach (Misgurnus anguillicaudatus) Protein Hydrolysates. Innovative Food Science & Emerging Technologies, 10(2), 235-240.
Bamdad, F., Wu, J. & Chen, L. (2011). Effects of Enzymatic Hydrolysis on Molecular Structure and Antioxidant Activity of Barley Hordein. Journal of Cereal Science, 54, 20-28.
Bondet, V., Brand-Williams, W. & Berset, C. (1997). Kinetics and Mechanisms of Antioxidant Activity Using the DPPH. Free Radical Method. LWT-Food Science and Technology, 30(6), 609-615.
Chiang, W. D., Shih, C. J. & Chu, Y. H. (1999). Functional Properties of Soy Protein Hydrolysate Produced from a Continuous Membrane Reactor System. Food Chemistry, 65(2), 189-194.
Dryáková, A., Pihlanto, A., Marnila, P., Čurda, L. & Korhonen, H. J. (2010). Antioxidant Properties of Whey Protein Hydrolysates as Measured by Three Methods. European Food Research and Technology, 23 (6), 865- 874.
Fitznar, H. P., Lobbes, J. M. & Kattner, G. (1999). Determination of Enantiomeric Amino acids with High-Performance Liquid Chromatography and Pre-column Derivatisation with O-phthaldialdehyde and N-Isobutyrylcysteine in Seawater and Fossil Samples (mollusks). Journal of Chromatography, A832(1), 123-132.
He, R., Alashi, A., Malomo, S. A., Girgih, A. T., Cha D., Ju, X. & Aluko, R. E. (2013). Antihypertensive and Free Radical Scavenging Properties of Enzymatic Rapeseed Protein Hydrolysates. Food Chemistry, 141(1), 153-159.
Hou, R. Z. Y., Yang, G. Li, Y. B., Huang, H., Wang, Y. J., Liu, L. & Zhang, X. Z. (2006). Synthesis of a Precursor Dipeptide of RGDS (Arg‐Gly‐Asp‐Ser) Catalysed by the Industrial Protease Alcalase. Biotechnology and Applied Biochemistry, 44(2), 73-80.
Lowry, O., Rosebrough, N., Farr, A. & Randall, R. (1951). Protein Estimation by Lowry’s Method. The Journal of Biological Chemistry, 193(1), 265-275.
Ludescher, R. D. (1996). Physical and Chemical Properties of Amino Acids and Proteins. Food Proteins: Properties and Characterization: Wiley-VCH, PP. 23-70.
Makinen, S. (2014). Production, Isolation and Characterization of Bioactive Peptides with Antihypertensive Properties from Rapeseed and Potato Protein, phD thesis, University of turku.
Martysiak-Zurowska, D. & Wenta, W. (2012). A Comparison of ABTS and DPPH Methods for Assessing the Total Antioxidant Capacity of Human Milk. Acta Scientiarum Polonorum. Technologia Alimentaria, 11(1), 83-89.
Moure, A., Domínguez, H. & Parajó, J. C. (2006). Antioxidant Properties of Ultrafiltration-Recovered Soy Protein Fractions from Industrial Effluents and Their Hydrolysates. Process Biochemistry, 41(2), 447-456.
Morr, C. V., German, B., Kinsella J. E., Regenstein, J., Van Buren, J. P., Kilara, A., Lewis, B. A. & Mangino, M. E. (1985). A Collaborative Study to Develop a Standardized Food Protein Solubility Procedure. Journal of Food Science 50(6): 1715-1718.
Nwokolo, E. & Smartt, J. (1996). Food and Feed from Legumes and Oilseeds, Chapman & Hall London, UK.
Panyam, D. & Kilara, A. (1996). Enhancing the Functionality of Food Proteins by Enzymatic Modification. Trends in Food Science & Technology, 7(4), 120-125.
Quist, E. E. (2005). Peanut (Arachis hypogaea L.) as a Sourse of Antihypertensive and Antimicrobial Peptides. MSc thesis, University of Ghan.
Re, R., Pellegrini, N., Proteggente, A., Pannala, A., Yang, M. & .Rice-Evans, C. (1999). Antioxidant Activity Applying an Improved ABTS Radical Cation Decolorization Assay. Free Radical Biology and Medicine, 26(9), 1231-1237.
Saito, Y., Wanezaki, K., Kawato, A. & Imayasu, S. (1994). Structure and Activity of Angiotensin I Converting Enzyme Inhibitory Peptides from Sake and Sake Lees. Bioscience, Biotechnology, and Biochemistry, 58(10), 1767-1771.
Salunkhe,, D. K. (1992). World Oilseeds, Springer Science & Business Media.
Siow, H. L. & Gan, C. Y. (2013). Extraction of Antioxidative and Antihypertensive Bioactive Peptides from Parkia speciosa Seeds. Food Biochemistry, 141, 3435–3442.
Taheri, A., Jalalinezhad, S. & Anvar, S. A. A. (2012). Antihypertensive and Antioxidant Properties of Five Different Protein Hydrolysates Produced from Indian White Shrimp (Penaeus indicus) By-products. Journal of Comparative Pathobiology, 9 (1), 599-608 [In Persian].
Tang, C. H., Wang, X. S. & Yang, X. Q. (2009). Enzymatic Hydrolysis of Hemp (Cannabis sativa L.) Protein Isolate by Various Proteases and Antioxidant Properties of the Resulting Hydrolysates. Food Chemistry, 141, 1484–1490.
Turner, J. & Backman, P. (1991). Factors Relating to Peanut Yield Increases after Seed Treatment with Bacillus Subtilis. Plant Disease, 75(4): 347-353.
Wieser, H. (2007). Chemistry of Gluten Proteins. Food Microbiology, 24(2), 115-119.
Woodroof, J. (1983). Peanut Butter. Peanuts: Production, Processing and Products, AVI Publishing co, Inc, 3rd edn, West port, Connecticut 127.
You, L., Zhao, M., Cui, C., Zhao, H. & Yang, B. (2009). Effect of Degree of Hydrolysis on the Antioxidant Activity of Loach (Misgurnus anguillicaudatus) Protein Hydrolysates. Innovative Food Science & Emerging Technologies, 10(2), 235-240.