Document Type : Full Research Paper


Department of Food Science and Technology, Faculty of Agriculture, Ferdowsi University of Mashhad, Mashhad, Iran.


Introduction: Lipid oxidation leads to the generation of off-flavors and potential toxic compounds. Synthetic antioxidants are frequently applied for inhibiting this reaction, however; there is a concern regarding to the potent toxic effects of synthetic antioxidants on human health. The non-enzymatic glycosylation reaction (Maillard reaction) has been broadly used to ameliorate the biological and functional features of proteins and polysaccharides. The Maillard reaction produces products with versatile functions such as antioxidant, antimicrobial, antihypertensive, anti-browning, and prebiotic properties. In this regard, the Maillard reaction products (MRPs) can be used in the food industry to inhibit the oxidation reaction due to their superb antioxidant effect. In this study, chitosan was glycosylated with inulin, fructose, and glucose. Chitosan is a chitin derivative with cationic nature having antimicrobial, antioxidant, metal chelation, and film-forming features. Inulin is recognized as a prebiotic sugar with vast applications in food and pharmaceutical sciences. The purpose of this study was to chemically modify chitosan through the Maillard reaction in order to boost its antioxidant and antimicrobial properties.
Materials and methods: Chitosan (0.5% w/v) was dissolved in 1.0% v/v acetic acid solution followed by stirring for 1.0 h at room temperature. Afterwards, sugars inulin, glucose, and fructose were separately added to the chitosan solution at final concentration of 1.0% w/v. The obtained solutions were then stirred until complete sugar dissolution. The pH of solution was adjusted to 6.07 by adding 2.0 M sodium hydroxide and then the chitosan-sugar Maillard conjugates were fabricated through autoclaving the solutions at 121 °C. Changes in pH after the reaction were measured using a pH meter. The extent of the Maillard reaction was estimated via measuring the absorbance of the conjugated solutions at 294 nm (the intermediate products) and 420 nm (final products). Fourier transform infrared (FTIR) spectroscopy at transmission mode and 400-4000 cm-1 was employed to evaluate the structural changes of chitosan upon conjugation. Antioxidant activity of the conjugates was evaluated based on the reducing power assay. One mL of the samples was charged with 1.0 mL of distilled water and 1.0 mL of potassium ferricyanide (1.0% w/v). The solution was mixed and incubated at 50 °C for 20 min. After adding 2.5 mL of tri-chloroacetic solution (10% w/v), the obtained solution was centrifuged at 5000 g for 5.0 min. Afterwards, 2.0 mL of the supernatant was mixed with 2.0 mL of distilled water and 1.0 mL of ferric chloride (0.1% w/v). The solution was stand for 10 min at ambient temperature and then its absorbance was recorded at 700 nm. Antimicrobial effect of the conjugates against pathogenic microorganisms (E. coli, S. aureus, B. subtilis, P. aeruginosa, A. niger, and C. albicans) was measured according to the minimum inhibitory (MIC) and microbiocidal (MBC) concentrations. SPSS software (version 21) and one-way ANOVA were applied for data analysis. Duncan’s multiple range test was employed to determine the differences between means.
Results & discussion: The Maillard reaction led to a significant decrement in pH value of chitosan-saccharide systems, mainly due to the covalent coupling of amino groups of chitosan to carbonyl groups of reducing sugars in conjugation with the production of acetic and formic acids. The highest intermediate compounds (A 294nm) and lowest browning intensity (A 420nm) observed in chitosan-fructose conjugate, which was likely attributed to the lower reactivity of fructose. Chitosan-inulin conjugate presented the highest A 420nm and lowest intermediate-to-final ratio (A 294nm/A 420nm), probably due to the lower inulin molecules and subsequently carbonyl groups compared to fructose and glucose. These groups may react with amino groups of chitosan at initial reaction times, leading more conversion rate of the intermediate compounds to the final ones. FTIR spectra of the chitosan and conjugates revealed that absorbance peak at 1661 cm-1 in chitosan spectrum decreased and shifted to 1578 cm-1 (in chitosan-fructose conjugate), 1579 cm-1 (in chitosan-glucose conjugate), and 1580 cm-1 (in chitosan-inulin conjugate), indicating the stretching C-N group and -C=N group and the formation of Schiff base (-C=N) between reducing end of the saccharides and amino groups of chitosan. Reducing power of the chitosan-saccharide systems improved after the thermal process. Although, chitosan-glucose and chitosan-fructose conjugates had significantly higher reducing power than unconjugated counterparts, but chitosan-inulin conjugate showed non-significantly improved antioxidant activity compared to its non-heated mixture. Antioxidant activity of the Maillard conjugates was ascribed from the electron donating ability of their hydroxyl and pyrrole groups. The conjugates had lower MIC and MBC in comparison to their unconjugated pairs, except for chitosan-glucose conjugate, which showed no differences in MIC and MBC compared with its non-heated mixture. Antimicrobial property of the Maillard products, especially melanoidins has been attributed to their metal chelating features; melanoidins exert a bacteriostatic effect at low concentration and bactericidal effect at high levels through sequestering ionic iron from medium and magnesium from outer membrane, leading to the cell membranes destabilization. Additionally, antioxidant capacity, high surface activity, and inhibiting effect towards catabolic enzymes have been reported as another antimicrobial mechanisms of the Maillard products. In general, it can be concluded that chitosan-saccharide Maillard-based conjugates, particularly inulin-chitosan one could be used in the food sector as a novel prebiotic-based active bio-compound with antioxidant and antimicrobial features.


Akagawa, M., Sasaki, T., & Suyama, K., 2002, Oxidative deamination of lysine residue in plasma protein of diabetic rats. The FEBS Journal, 269(22), 5451-5458.
Behbahani, B. A., Shahidi, F., Yazdi, F. T., Mortazavi, S. A., & Mohebbi, M., 2017, Antioxidant activity and antimicrobial effect of tarragon (Artemisia dracunculus) extract and chemical composition of its essential oil. Journal of Food Measurement and Characterization, 11(2), 847-863.
Benjakul, S., Lertittikul, W., & Bauer, F., 2005, Antioxidant activity of Maillard reaction products from a porcine plasma protein–sugar model system. Food Chemistry, 93(2), 189-196.
Beverlya, R. L., Janes, M. E., Prinyawiwatkula, W., & No, H. K., 2008, Edible chitosan films on ready-to-eat roast beef for the control of Listeria monocytogenes. Food Microbiology, 25(3), 534-537.
Chang, H. L., Chen, Y. C., & Tan, F. J., 2011, Antioxidative properties of a chitosan–glucose Maillard reaction product and its effect on pork qualities during refrigerated storage. Food chemistry, 124(2), 589-595.
de Oliveira, F. C., Coimbra, J. S. D. R., de Oliveira, E. B., Zuñiga, A. D. G., & Rojas, E. E. G., 2016, Food protein-polysaccharide conjugates obtained via the maillard reaction: A review. Critical Reviews in Food Science and Nutrition, 56(7), 1108-1125.
Devlieghere, F., Vermeulen, A., & Debevere, J., 2004, Chitosan: antimicrobial activity, interactions with food components and applicability as a coating on fruit and vegetables. Food microbiology, 21(6), 703-714.
Gullon, B., Montenegro, M. I., Ruiz-Matute, A. I., Cardelle-Cobas, A., Corzo, N., & Pintado, M. E., 2016, Synthesis, optimization and structural characterization of a chitosan–glucose derivative obtained by the Maillard reaction. Carbohydrate polymers, 137, 382-389.
He, Y., 2015, Improved heat stability of whey protein isolate by glycation with inulin. University of Missouri-Columbia.
Jiang, Z., Rai, D. K., O'Connor, P. M., & Brodkorb, A., 2013, Heat-induced Maillard reaction of the tripeptide IPP and ribose: Structural characterization and implication on bioactivity. Food research international, 50(1), 266-274.
Jing, H., Yap, M., Wong, P. Y., & Kitts, D. D., 2011, Comparison of physicochemical and antioxidant properties of egg-white proteins and fructose and inulin Maillard reaction products. Food and Bioprocess Technology, 4(8), 1489-1496.
Kalyani Nair, K., Kharb, S., & Thompkinson, D. K., 2010, Inulin dietary fiber with functional and health attributes—a review. Food Reviews International, 26(2), 189-203.
Karimi, R., Azizi, M. H., Ghasemlou, M., & Vaziri, M., 2015, Application of inulin in cheese as prebiotic, fat replacer and texturizer: A review. Carbohydrate Polymers, 119, 85-100.
Kim, J. S., & Lee, Y. S., 2009, Study of Maillard reaction products derived from aqueous model systems with different peptide chain lengths. Food Chemistry, 116(4), 846-853.
Kumar, M. N. R., 2000, A review of chitin and chitosan applications. Reactive and functional polymers, 46(1), 1-27.
Labuza, T. P., Monnier, V., Baynes, J., & O'Brien, J. (Eds.)., 1998, Maillard reactions in chemistry, food and health. Elsevier.
Li, X., Shi, X., Jin, Y., Ding, F., & Du, Y., 2013, Controllable antioxidative xylan–chitosan Maillard reaction products used for lipid food storage. Carbohydrate polymers, 91(1), 428-433.
Lingnert, H., & Eriksson, C. E., 1980, Antioxidative Maillard reaction products. II.
Products from sugars and peptides or protein hydrolysates. Journal of Food Processing
and Preservation, 4(3), 173-181.
Liu, S. C., Yang, D. J., Jin, S. Y., Hsu, C. H., & Chen, S. L., 2008, Kinetics of color development, pH decreasing, and anti-oxidative activity reduction of Maillard reaction in galactose/glycine model systems. Food Chemistry, 108(2), 533-541.
Lopes, S. M., Krausova, G., Rada, V., Gonçalves, J. E., Gonçalves, R. A., & de Oliveira, A. J., 2015, Isolation and characterization of inulin with a high degree of polymerization from roots of Stevia rebaudiana (Bert.) Bertoni. Carbohydrate research, 411, 15-21.
Martins, S. I., Jongen, W. M., & Van Boekel, M. A., 2000, A review of Maillard reaction in food and implications to kinetic modelling. Trends in Food Science & Technology, 11(9), 364-373.
Mcdevitt-Pugh, M., & Meyer, D., 2005, Low glycemic index products with inulin to support weight management. Wellness Foods Europe, 34, 20-24.
Muppalla, S. R., Sonavale, R., Chawla, S. P., & Sharma, A., 2012, Functional properties of nisin–carbohydrate conjugates formed by radiation induced Maillard reaction. Radiation Physics and Chemistry, 81(12), 1917-1922.
Mutanda, T., Mokoena, M. P., Olaniran, A. O., Wilhelmi, B. S., & Whiteley, C. G., 2014, Microbial enzymatic production and applications of short-chain fructooligosaccharides and inulooligosaccharides: recent advances and current perspectives. Journal of industrial microbiology & biotechnology, 41(6), 893-906.
Nooshkam, M., & Madadlou, A., 2016a, Maillard conjugation of lactulose with potentially bioactive peptides. Food Chemistry, 192, 831-836.
Nooshkam, M., & Madadlou, A., 2016b, Microwave-assisted isomerisation of lactose to lactulose and Maillard conjugation of lactulose and lactose with whey proteins and peptides. Food chemistry, 200, 1-9.
Nursten, H. E., 2005, The Maillard reaction: chemistry, biochemistry, and implications.
Royal Society of Chemistry.
O'Brien, J., Morrissey, P. A., & Ames, J. M., 1989, Nutritional and toxicological aspects of the Maillard browning reaction in foods. Critical Reviews in Food Science & Nutrition, 28(3), 211-248.
Phisut, N., & Jiraporn, B., 2013, Characteristics and antioxidant activity of Maillard reaction products derived from chitosan-sugar solution. International Food Research Journal, 20(3).
Prashanth, K. H., & Tharanathan, R. N., 2007, Chitin/chitosan: modifications and their unlimited application potential—an overview. Trends in food science & technology, 18(3), 117-131.
Rufian-Henares, J. A., & de la Cueva, S. P., 2009, Antimicrobial Activity of Coffee Melanoidins A Study of Their Metal-Chelating Properties. Journal of Agricultural and Food Chemistry, 57(2), 432-438.
Rufian-Henares, J. A., & Morales, F. J., 2006, A new application of a commercial microtiter plate-based assay for assessing the antimicrobial activity of Maillard reaction products. Food Research International, 39(1), 33-39.
Rufian-Henares, J. A., & Morales, F. J., 2008a, Microtiter plate-based assay for screening antimicrobial activity of melanoidins against E. coli and S. aureus. Food chemistry, 111(4), 1069-1074.
Rufian-Henares, J. A., & Morales, F. J., 2008b, Antimicrobial activity of melanoidins against Escherichia coli is mediated by a membrane-damage mechanism. Journal of Agricultural and Food Chemistry, 56(7), 2357-2362.
Schaller‐Povolny, L. A., & Smith, D. E., 1999, Sensory attributes and storage life of reduced fat ice cream as related to inulin content. Journal of Food Science, 64(3), 555-559.
Shahidi, F., 2000, Antioxidants in food and food antioxidants. Food/nahrung, 44(3), 158-163.
Vhangani, L. N., & Van Wyk, J., 2013, Antioxidant activity of Maillard reaction products
(MRPs) derived from fructose–lysine and ribose–lysine model systems. Food Chemistry, 137(1), 92-98.
Wang, H. Y., Qian, H., & Yao, W. R., 2011, Melanoidins produced by the Maillard reaction: Structure and biological activity. Food Chemistry, 128(3), 573-584.
Wang, W. Q., Bao, Y. H., & Chen, Y., 2013, Characteristics and antioxidant activity of water-soluble Maillard reaction products from interactions in a whey protein isolate and sugars system. Food Chemistry, 139(1), 355-361.
Wu, S., Hu, J., Wei, L., Du, Y., Shi, X., & Zhang, L., 2014, Antioxidant and antimicrobial
activity of Maillard reaction products from xylan with
chitosan/chitooligomer/glucosamine hydrochloride/taurine model systems. Food Chemistry, 148, 196-203.
Yu, M., He, S., Tang, M., Zhang, Z., Zhu, Y., & Sun, H., 2018, Antioxidant activity and
sensory characteristics of Maillard reaction products derived from different peptide
fractions of soybean meal hydrolysate. Food Chemistry, 243, 249-257.
Yuan, D., Xu, Y., Wang, C., Li, Y., Li, F., Zhou, Y., ... & Jiang, Y., 2015, Comparison of anti-browning ability and characteristics of the fractionated Maillard reaction products with different polarities. Journal of Food Science and Technology, 52(11), 7163-7172.
Zhang, H., Yang, J., & Zhao, Y., 2015, High intensity ultrasound assisted heating to improve solubility, antioxidant and antibacterial properties of chitosan-fructose Maillard reaction products. LWT-Food Science and Technology, 60(1), 253-262.
Zhong, N. J., Liu, G. Q., Zhao, X. H., Gao, Y. Q., Li, L., & Li, B., 2015, Lipid Peroxidation Inhibitation Activity of Maillard Reaction Products Derived from Sugar-amino Acid Model Systems. Advance Journal of Food Science and Technology, 9(5), 393-397.