Antibacterial Activity and Characterization of Water-Soluble Chitosan Compounds Produced from Enzymatic Deacetylation

Yıl 2022, Cilt: 18 Sayı: 4, 451 - 460, 01.12.2022

Fatih Matyar Hatice Aysun Mercimek Takcı Fatma Yılmaz Gulcihan Guzeldag Halil İbrahim Çelik

https://doi.org/10.22392/actaquatr.1074431

Öz

In this research, it was aimed to investigate the production of water-soluble chitosan from the enzymatic deacetylation of chitin by Bacillus (B. cereus (BC) and B. thuringiensis (BT)) strains. Characteristic properties involving molecular weight, degree of deacetylation, and antibacterial activity of chitosan samples were identified. The degree of deacetylation of BC and BT chitosan samples obtained at 393 and 213 ppm amounts was reported as 80.54% and 86.07% by the IR spectrum. As well as the degree of deacetylation, the molecular weights of samples showed a significant effect on antibacterial activity were 48.27 and 48.46 Da, respectively. Among the tested bacteria, the highest inhibitory effect was recorded in Pseudomonas aeruginosa and Staphylococcus epidermis for both chitosan samples. On the other hand, no antibacterial effect on S. aureus, Vancomycin-resistant Enterococci, S. epidermidis, Klebsiella sp., and Salmonella spp. were observed. Our results indicated a simple and cost-efficient method for the production of chitosan polymers showing antibacterial activity. 

Anahtar Kelimeler

Antibacterial activity, Bacillus sp., chitosan, deacetylation, FTIR

Enzimatik Deasetilasyon ile Üretilen Su Çözülebilir Kitosan Bileşiklerinin Antibakteriyel Aktivitesi ve Karakterizasyonu

Öz

Bu araştırmada, Bacillus sp. (B. cereus (BC) ve B. thuringiensis (BT)) suşları ile kitinin enzimatik deasetilasyonundan suda çözülebilir kitosan üretiminin incelenmesi amaçlanmıştır. Kitosan örneklerinin moleküler ağırlıkları, deasetilasyon dereceleri ve antibakteriyel aktivitelerini içeren karakteristik özellikleri tanımlanmıştır. 393 (BC) ve 213 (BT) ppm miktarda elde edilen kitosan örneklerinin deasetilasyon derecesi IR spektrumu ile sırasıyla %80,54 ve %86,07 olarak belirlenmiştir. Deasetilasyon derecesinin yanı sıra antibakteriyel aktivitede önemli bir etki gösteren moleküler ağırlık değerleri ise 48,27 ve 48,46 Da’dır. Her iki kitosan bileşikleri için, test bakterileri arasında en yüksek inhibitör etki P. aeruginosa ve S. epidermis’e karşı kaydedilmiştir. Bunun yanı sıra S. aureus, Vankomisin dirençli Enterococci, S. epidermidis, Klebsiella sp. ve Salmonella spp’ye karşı hiçbir antibakteriyel etki gözlenmemiştir. Bizim sonuçlarımız, antibakteriyel aktivite gösteren kitosan polimerlerin üretimi için basit ve düşük maliyetli bir yönteme işaret etmektedir.

Anahtar Kelimeler

Antibakteriyel aktivite, Bacillus, kitosan, Deasetilasyon, FTIR

Kaynakça

• Abdeltwab, W. M., Abdelaliem, Y. F., Metry, W. A., Eldeghedy, M., & Ebid, W. (2019). Antimicrobial effect of chitosan and nano-chitosan against some pathogens and spoilage microorganisms. Journal of Advanced Laboratory Research in Biology, 10(1), 8-15.
• Ak Kalut, S., (2008). Enhancement of degree of deacetylation of chitin in chitosan production. [Bachelor thesis, University Malaysia Pahang].
• Ali, Z. H., & Aldujaili, N. H. (2022). Bio-Environmental preparation of chitosan nanoparticle using Bacillus subtilis and their biomedical activity. IOP Conf. Ser.: Earth Environmental Science, 1029, 012023. https://doi.org/10.1088/1755-1315/1029/1/012023
• Brasselet, C., Pierre, G., Dubessay, P., Dols-Lafargue, M., Coulon, J., Maupeu, J., Vallet-Courbin, A., De Baynast, H., Doco, T., Michaud, P., & Delattre, C. (2019). Modification of chitosan for the generation of functional derivatives. Applied Sciences, 9(7), 1321. https://doi.org/10.3390/app9071321
• Chung, Y. C., Yeh, J. Y., & Tsai, C. F. (2011). Antibacterial characteristics and activity of water-soluble chitosan derivatives prepared by the maillard reaction. Molecules, 16(10), 8504-8514. https://doi.org/10.3390/molecules16108504
• Dragland, I. S., Wellendorf, H., Kopperud, H., Stenhagen, I., & Valen, H. (2019). Investigation on the antimicrobial activity of chitosan-modified zinc oxide-eugenol cement. Biomaterial Investigations in Dentistry, 6(1), 99-106. https://doi.org/10.1080/26415275.2019.1697621
• Elmekawy, A., Hegab, H. M., El-Baz, A., & Hudson, S. M. (2013). Kinetic properties and role of bacterial chitin deacetylase in the bioconversion of chitin to chitosan. Recent Patents on Biotechnology, 7(3), 234-241. https://doi.org/10.2174/1872208307666131202192453
• Goy, R. C., Britto, D., & Assis, O. B. G. (2009). A Review of the antimicrobial activity of chitosan. Polímeros, 19(3), 241-247. https://doi.org/10.1590/S0104-14282009000300013
• Junior, J. C. V., Ribeaux, D. R., Alves Da Silva, C. A., & Campos-Takakı, G. M. D. (2016). Physicochemical and antibacterial properties of chitosan extracted from waste shrimp shells. International Journal of Microbiology, 2016, ID: 5127515. https://doi.org/10.1155/2016/5127515
• Kasaai, M. R. (2008). A review of several reported procedures to determine the degree of N-acetylation for chitin and chitosan using infrared spectroscopy. Carbohydrate Polymers, 71(4), 497-508. https://doi.org/10.1016/j.carbpol.2007.07.009
• Kaur, K., Dattajirao, V., Shrivastava, V., & Bhardwaj, U. (2012). Isolation and characterization of chitosan-producing bacteria from beaches of Chennai, India. Enzyme Research, 2012(4), 421683. https://doi.org/10.1155/2012/421683
• Kaczmarek, M. B., Struszczyk-Swita, K., Li, X., Szczesna-Antczak, M., & Daroch, M. (2019). Enzymatic modifications of chitin, chitosan and chitooligosaccharides. Frontiers in Bioengineering and Biotechnology, 7, 243. https://doi.org/10.3389/fbioe.2019.00243
• Leane, M. M., Nankervis, R., Smith, R., & Illum, L. (2004). Use of the ninhydrin assay to measure the release of chitosan from oral solid dosage forms. International Journal of Pharmaceutics, 271(1-2), 241-249. https://doi.org/10.1016/j.ijpharm.2003.11.023
• Mercimek Takci, H. A., Karayilan, R., Ucan Turkmen, F., Sumengen Ozdenefe, M., Buyukkaya Kayis, F., Arkut Ozyapıcı, A., & Caktu Guler, K. (2019). Biochemical characterization of the crude chitin deacetylase supernatant Bacillus cereus. Fresenius Environmental Bulletin, 28(9), 6661-6667.
• Mohammadi, B., Pirsa, S., & Alizadeh, M. B. (2019). Preparing chitosan–polyaniline nanocomposite film and examining its mechanical, electrical, and antimicrobial properties. Polymers & Polymer Composites, 27(8), 507-517. https://doi.org/10.1177/0967391119851439
• Ortega-Ortiz, H., Gutiérrez-Rodríguez, B., Cadenas-Pliego, G., & Jimenez, L. (2010). Antibacterial activity of chitosan and the interpolyelectrolyte complexes of poly(acrylic acid)- chitosan. Brazilian Archives of Biology and Technology, 53(3), 623-628. https://doi.org/10.1590/S1516-89132010000300016
• Pareek, N., Vivekanand, V., Agarwal, P., Saroj, S., & Singh, R. P. (2013). Bioconversion to chitosan: A two stage process employing chitin deacetylase from Penicillium oxalicum SAEM-51. Carbohydrate Polymers, 96(2), 417-425. https://doi.org/10.1016/j.carbpol.2013.04.005
• Prabu, K., & Nataraja, E. (2012). In vitro antimicrobial and antioxidant activity of chitosan isolated from Podophthalmus vigil. Journal of Applied Pharmaceutical Science, 2(9), 75-82. https://doi.org/10.7324/JAPS.2012.2916
• Prochazkova, S., Vårum, K. M., & Østgaard, K. (1999). Quantitative determination of chitosans by ninhydrin. Carbohydrate Polymers, 38(2), 115-122. https://doi.org/10.1016/S0144-8617(98)00108-8
• Qin, Y., & Li, P. (2020). Antimicrobial chitosan conjugates: current synthetic strategies and potential applications. International Journal of Molecular Sciences, 21(2), 499. https://doi.org/10.3390/ijms21020499
• Saito, H., Sakakibara, Y., Sakata, A., Kurashige, R., Murakami, D., Kageshima, H., Saito, A., & Miyazaki, Y.. (2019). Antibacterial activity of lysozyme-chitosan oligosaccharide conjugates (LYZOX) against Pseudomonas aeruginosa, Acinetobacter baumannii and Methicillin-resistant Staphylococcus aureus. Plos One, 14(5), e0217504. https://doi.org/10.1371/journal.pone.0217504
• Sharma, R., Raghav, R., Priyanka, K., Rishi, P., Sharma, S., Srivastava, S., & Verma, I. (2019). Exploiting chitosan and gold nanoparticles for antimycobacterial activity of in silico identified antimicrobial motif of human neutrophil peptide-1. Scientific Reports, 9(1), 7866. (2019) 9:7866. https://doi.org/10.1038/s41598-019-44256-6
• Sahariah, P., & Masson, M. (2017). Antimicrobial chitosan and chitosan derivatives: a review of the structure–activity relationship. Biomacromolecules, 18(11), 3846-3868. https://doi.org/10.1021/acs.biomac.7b01058
• Santos, V. P., Marques, N. S., Maia, P. C., Lima, M. A. B. D., Franco, L. D. O., & Campos-Takaki, G. M. D. (2020). Seafood waste as attractive source of chitin and chitosan production and their applications. International Journal of Molecular Sciences, 21(12), 4290. https://doi.org/10.3390/ijms21124290
• Tuveng, T. R., Rothweiler, U., Udatha, G., Vaaje-Kolstad, G., Smalås, A., & Eijsink, V. G. H. (2017). Structure and function of a CE4 deacetylase isolated from a marine environment. PLoS One, 12(11), e0187544. https://doi.org/10.1371/journal.pone.0187544
• Venugopal, V. (2021). Valorization of seafood processing discards: Bioconversion and bio-refinery approaches. Frontiers in Sustainable Food Systems, 132. https://doi.org/10.3389/fsufs.2021.611835
• Yomota, C., Miyazaki, T., & Okada, S. (1993). Determination of the viscometric constants for chitosan and the application of universal calibration procedure in its gel permeation chromatography. Colloid and Polymer Science, 271, 76-82. https://doi.org/10.1016/0141-8130(82)90074-5
• Yonis, R. W., Luti, K. J., & Aziz, G. M. (2019). Statistical optimization of chitin bioconversion to produce an effective chitosan in solid state fermentation by Asperigellus flavus. Iraqi Journal of Agricultural Sciences, 50(3), 916-927. https://doi.org/10.36103/ijas.v50i3.708