The Cytotoxicity Effect of Agaro-Oligosaccharides and Neoagaro-Oligosaccharides from Different Enzymatic Hydrolysis Time on Macrophage Cells

Visi Endah Pratitis, Pinki Anggrahini Puspitasari, Mayland Muhammad Hanbal, Khansa Tsabitah, Dian Juliadmi, Budi Saksono, Nastiti Wijayanti

Abstract


Agarose is a polysaccharide from red algae. Enzymatic hydrolysis of agarose can produce Agaro-Oligosaccharidess (AOS) and Neoagaro-Oligosaccharidess (NAOS). Different times of enzymatic hydrolysis can produce different types of AOS and NAOS based on their degree of polymerization (DPs). This study aims to examine the cytotoxicity of AOS and NAOS with different hydrolysis times on the macrophage cell line RAW 264.7. The parameters used were the percentage of cell viability and IC50 value. The cytotoxicity test using MTT assay and One-way ANOVA were used as statistical tests. The results showed that AOS-0 at a concentration below 125 µg/mL was not toxic and showed moderate toxicity up to a high concentration of 1000 µg/mL, while AOS hydrolyzed for 24 h (AOS-24) was not toxic to RAW 264.7 cells at all concentrations tested. The different results were shown in all NAOS samples, which were highly toxic to RAW 264.7 cells in the 125 to 1000 µg/mL, indicating that it was concentration dependent. The results showed that different hydrolysis times caused differences in the structure of AOS and NAOS compounds and influenced the toxicity level. Research development for further studies on antioxidants and anti-inflammatory needs more attention to the sample type and hydrolysis time.

Keywords


AOS; NAOS; Enzymatic hydrolysis; Cytotoxicity

References


Usov AI. Polysaccharides of the red algae [Internet]. 1st ed. Vol. 65, Advances in Carbohydrate Chemistry and Biochemistry. Elsevier Inc.; 2011. 115–217 p. http://dx.doi.org/10.1016/B978-0-12-385520-6.00004-2

De Castro JPL, Costa LEC, Pinheiro MP, Dos Santos Francisco T, De Vasconcelos PHM, Funari LM, et al. Polysaccharides of red alga Gracilaria intermedia: Structure, antioxidant activity and rheological behavior. Polimeros. 2018;28(2):178–86. https://doi.org/10.1590/0104-1428.013116

Pandey SP, Shukla T, Dhote VK, Mishra DK, Maheshwari R, Tekade RK. Use of Polymers in Controlled Release of Active Agents [Internet]. Basic Fundamentals of Drug Delivery. Elsevier Inc.; 2019. 113–172 p. http://dx.doi.org/10.1016/B978-0-12-817909-3.00004-2

Cheong KL, Qiu HM, Du H, Liu Y, Khan BM. Oligosaccharides derived from red seaweed: Production, properties, and potential health and cosmetic applications. Molecules. 2018;23(10). https://doi.org/10.3390/molecules23102451

Yun EJ, Lee S, Kim JH, Kim BB, Kim HT, Lee SH, et al. Enzymatic production of 3,6-anhydro-l-galactose from agarose and its purification and in vitro skin whitening and anti-inflammatory activities. Appl Microbiol Biotechnol. 2013;97(7):2961–70. https://doi.org/10.1007/s00253-012-4184-z

Yun EJ, Yu S, Kim KH. Current knowledge on agarolytic enzymes and the industrial potential of agar-derived sugars. Appl Microbiol Biotechnol. 2017;101(14):5581–9. https://doi.org/10.1007/s00253-017-8383-5

Kim DH, Park SY, Kim KH. Optimal β-galactosidases for producing high-titer 3,6-anhydro-L-galactose from red-algal agarobiose. Appl Microbiol Biotechnol [Internet]. 2022;106(24):8111–20. https://doi.org/10.1007/s00253-022-12274-6

Hong SJ, Lee JH, Kim EJ, Yang HJ, Chang YK, Park JS, et al. In vitro and in vivo investigation for biological activities of neoagarooligosaccharides prepared by hydrolyzing agar with β-agarase. Biotechnol Bioprocess Eng. 2017;22(4):489–96. https://doi.org/10.1007/s12257-017-0049-8

Qu W, Wang D, Wu J, Chan Z, Di W, Wang J, et al. Production of Neoagaro-Oligosaccharides With Various Degrees of Polymerization by Using a Truncated Marine Agarase. Front Microbiol. 2020;11(September). https://doi.org/10.3389/fmicb.2020.574771

Zhang YH, Song XN, Lin Y, Xiao Q, Du XP, Chen YH, et al. Antioxidant capacity and prebiotic effects of Gracilaria neoagaro oligosaccharides prepared by agarase hydrolysis. Int J Biol Macromol [Internet]. 2019;137:177–86. https://doi.org/10.1016/j.ijbiomac.2019.06.20711.

Adan A, Kiraz Y, Baran Y. Cell Proliferation and Cytotoxicity Assays. Curr Pharm Biotechnol. 2016;17(14):1213. 10.2174/1389201017666160808160513

Van Tonder A, Joubert AM, Cromarty AD. Limitations of the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay when compared to three commonly used cell enumeration assays. BMC Res Notes. 2015;8(1):1–10. https://doi.org/10.1186/s13104-015-1000-8

Ghasemi M, Turnbull T, Sebastian S, Kempson I. The mtt assay: Utility, limitations, pitfalls, and interpretation in bulk and single-cell analysis. Int J Mol Sci. 2021;22(23). https://doi.org/10.3390/ijms222312827.

Bahuguna A, Khan I, Bajpai VK, Kang SC. MTT assay to evaluate the cytotoxic potential of a drug. Bangladesh J Pharmacol. 2017;12(2):115–8. https://doi.org/10.3329/bjp.v12i2.30892

Xu SY, Kan J, Hu Z, Liu Y, Du H, Pang GC, et al. Quantification of neoagaro-oligosaccharide production through enzymatic hydrolysis and its anti-oxidant activities. Molecules. 2018;23(6). https://doi.org/10.3390/molecules23061354

Chen HM, Yan XJ. Antioxidant activities of agaro-oligosaccharides with different degrees of polymerization in cell-based system. Biochim Biophys Acta - Gen Subj. 2005;1722(1):103–11. https://doi.org/10.1016/j.bbagen.2004.11.016.

Sujana PKW, Wijayanti N. Phytochemical and antioxidant properties of Syzygium zollingerianum leaves extract. Biodiversitas. 2022;23(2):916–21. https://doi.org/10.13057/biodiv/d230233.

Van den Berg BM. Microscopic analysis of MTT stained boar sperm cells. Open Vet J. 2015;5(1):58–63. http://dx.doi.org/10.13140/RG.2.1.4909.0726.

Susanty A, Febriana M, Putri DS, Ikhtiarudin I, Wahyuni FS, Dachriyanus. Cytotoxic Activity of Ethyl Acetate Extract from Voacanga foetida ( Bl .) Rolfe Leaves Against T47D Breast Cancer Cells. Borneo J Pharm. 2023;6(1):8–14. https://doi.org/10.33084/bjop.v6i1.3499.

Lee CH, Kim HT, Yun EJ, Lee AR, Kim SR, Kim JH, et al. A novel agarolytic β-galactosidase acts on agarooligosaccharides for complete hydrolysis of agarose into monomers. Appl Environ Microbiol. 2014;80(19):5965–73. https://doi.org/10.1128/AEM.01577-14

Kim JH, Yun EJ, Yu S, Kim KH, Kang NJ. Different levels of skin whitening activity among 3,6-anhydro-L-galactose, agarooligosaccharides, and neoagarooligosaccharides. Mar Drugs. 2017;15(10). https://doi.org/10.3390/md15100321.

Zou Y, Fu X, Liu N, Duan D, Wang X, Xu J, et al. The synergistic anti-inflammatory activities of agaro-oligosaccharides with different degrees of polymerization. J Appl Phycol. 2019;31(4):2547–58. https://doi.org/10.1007/s10811-019-1740-2




DOI: https://doi.org/10.18196/mmjkk.v24i2.21059

Refbacks

  • There are currently no refbacks.


Creative Commons License
This work is licensed under a Creative Commons Attribution 4.0 International License.

      

Editorial Office:
Journal Room, G1 (Biomedic) Building, Ground Floor, Faculty of Medicine and Health Science Universitas Muhammadiyah Yogyakarta, 
Jalan Lingkar Selatan (Brawijaya), Tamantirto, Kasihan, Bantul, Daerah Istimewa Yogyakarta, Indonesia
Phone: +62 274 387 656 (ext: 231)
WA : +62 811-2650-303
Website: http://journal.umy.ac.id/index.php/mm 
E-mail: mmjkk@umy.university

Creative Commons License
Mutiara Medika: Jurnal Kedokteran dan Kesehatan is licensed under a Creative Commons Attribution 4.0 International License. View My Stats