Putrescine의 존재 하에 중성 pH 조건에서의 가교 히알루론산 하이드로젤의 합성 및 특성

제유림; 방소라; 권일근; 박상준; 김천호; Yu Rim Je; So Ra Bang; Il Keun Kwon; Sang Jun Park; Chun-Ho Kim

Polymer(Korea), Vol.45, No.4, 601-609, July, 2021

DOI10.7317/pk.2021.45.4.601 Export Citation

Putrescine의 존재 하에 중성 pH 조건에서의 가교 히알루론산 하이드로젤의 합성 및 특성

Synthesis and Characteristic of Cross-linked Hyaluronic Acid Hydrogels with Putrescine under the Neutral pH Condition

제유림^{1, 2}, 방소라¹, 권일근², 박상준¹, 김천호^{1, †}

¹한국원자력의학원 방사선의학연구소
²경희대학교 치과대학 치의학과

Yu Rim Je^{1, 2}, So Ra Bang¹, Il Keun Kwon², Sang Jun Park¹, and Chun-Ho Kim^{1, †}

¹Laboratory of Tissue Engineering, Korea Institute of Radiological Medical Sciences, Seoul 01812, Korea
²Department of Dentistry, Graduate School, Kyung Hee University, 26 Kyungheedae-ro, Dongdaemum-gu, Seoul 02447, Korea

E-mail:

초록

본 연구는 히알루론산의 가교 반응 중 분해를 최소화하기 위해, 중성 pH 조건에서 히알루론산 하이드로젤을 제조하는 것이다. 가교 반응은 diamine인 putrescine의 존재 하에, 4-(4,6-dimethoxy-1,3,5-triazin-2-yl)-4-methyl-morpholinium chloride(DMTMM) 및 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide(EDC)/N-hydroxysuccinimide(NHS)를 각각 가교제로 사용하여 2종류의 히알루론산 하이드로젤을 제조하였다(HPDM 및 HPEN). FTIR 분석을 통해 두 가교제 모두 putrescine의 존재 하에, 아마이드기를 형성했음을 확인하였다. 반응에 따른 하이드로젤의 형태 및 유변학적 특성을 주사전자현미경 및 레오미터를 통해 확인하였다. 제조된 하이드로젤의 팽윤 특성 및 분해 거동은 가교제의 종류 및 putrescine/가교제의 몰비에 직접적인 영향을 받았다. 하이드로젤에서의 인체 섬유아세포에 대한 세포 적합성은 MTTassay를 통해 확인하였다. 결론적으로, DMTMM으로 가교한 HPDM의 가교 효율 및 생체 적합성이 EDC/NHS으로 가교한 HPEN 보다 더 우수하였다.

This study is to fabricate hyaluronic acid (HyA) hydrogels under the neutral pH condition in order to minimize the degradation of HyA during the cross-linking reaction. Two kinds of HyA hydrogels (HPDM or HPEN), were prepared with 4-(4,6-dimethoxy-1,3,5-triazin-2-yl)-4-methyl-morpholinium chloride (DMTMM) or 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC)/N-hydroxysuccinimide (NHS) as cross-linking agents, respectively, in the presence of putrescine as a diamine. Amide bond formation of hydrogels in the presence of putrescine was confirmed by FT-IR analysis. The morphology and rheological property of the hydrogels were studied through a scanning electron microscope and a rheometer, respectively. The swelling property and the degradation behavior of the hydrogels were affected by the two kinds of cross-linking agents and the molar ratio of the putrescine/cross-linking agents. Cytocompatibility of these hydrogels on human dermal fibroblast was confirmed by MTT assay. In conclusion, the cross-linking efficiency and the biocompatibility of HPDM cross-linked by DMTMM were better than those of HPEN cross-linked by EDC/NHS.

Keywords:neutral pH condition;putrescine;hyaluronic acid;4-(4,6-dimethoxy-1,3,5-triazin-2-yl)-4-methyl-morpholinium chloride;1-ethyl-3-(3-dimethylaminopropyl) carbodiimide/N-hydroxysuccinimide

[References]

Collins MN, Birkinshaw C, Carbohydr. Polym., 92, 1262 (2013)
Jung SW, Oh SH, Lee IS, Byun JH, Lee JH, Tissue Eng. Regen. Med., 16, 479 (2019)
Im GI, Tissue Eng. Regen. Med., 1 2019.
Lyu HZ, Lee JH, Biomaterials Res., 24, 5 (2020)
Choi JS, Oh SH, Kim YM, Lim JY, Tissue Eng. Regen. Med., 17, 651 (2020)
Min SJ, Kim SW, Lee TH, Kwon IK, Son TI, Polym. Korea, 44(4), 451 (2020)
Sung YK, Kim SW, Biomaterials Res., 24, 1 (2020)
Yang JM, Kim HJ, Cho BK, Polym. Korea, 44, 21 (2020)
Hong BM, Park SA, Park WH, Biomaterials Res., 23, 1 (2019)
Xu X, Jha AK, Harrington DA, Farach-Carson MC, Jia X, Soft Mater., 8, 3280 (2012)
Khunmanee S, Jeong Y, Park H, J. Tissue Eng., 8, 2041731417726464 2017.
Yang B, Guo X, Zang H, Liu J, Carbohydr. Polym., 131, 233 (2015)
Shimojo AA, Pires A, Lichy R, Santana MH, J. Brazil. Chem. Soc, 26, 506 (2015)
Ibrahim S, Kang QK, Ramamurthi A, J. Biomed. Mater. Res. A, 94, 355 (2010)
Calles JA, Ressia JA, Llabot JM, Valles EM, Palma SD, Sci. Pharm., 84, 61 (2016)
Kirk JF, Ritter G, Finger I, Sankar D, Reddy JD, Talton JD, et al>, Biomatter, 3, e25633 (2013)
Cao W, Sui J, Ma M, Xu Y, Lin W, Chen Y, Man Y, Sun Y, Fan Y, Zhang X, J. Mater. Chem. B, 7, 4413 (2019)
Nakajima N, Ikada Y, Bioconjugate Chem., 6, 123 (1995)
Bulpitt P, Aeschlimann D, J. Biomed. Mater. Res. A, 47, 152 (1999)
Kunishima M, Kawachi C, Monta J, Terao K, Iwasaki F, Tani S, Tetrahedron, 55, 13159 (1999)
Farkas P, Bystricky S, Carbohydr. Polym., 68, 187 (2007)
Raw SA, Tetrahedron Lett., 50, 946 (2009)
D’este M, Eglin D, Alini M, Carbohydr. Polym., 108, 239 (2014)
Til H, Falke H, Prinsen M, Willems M, Food Chem. Toxicol., 35, 337 (1997)
Farriol M, Segovia-Silvestre T, Castellanos JM, Venereo Y, Orta X, Nutrition, 17, 934 (2001)
Seliktar D, Science, 336(6085), 1124 (2012)
Tomihata K, Ikada Y, J. Biomed. Mater. Res. B, 37, 243 (1997)

[1] Collins MN, Birkinshaw C, Carbohydr. Polym., 92, 1262 (2013)

[2] Jung SW, Oh SH, Lee IS, Byun JH, Lee JH, Tissue Eng. Regen. Med., 16, 479 (2019)

[3] Im GI, Tissue Eng. Regen. Med., 1 2019.

[4] Lyu HZ, Lee JH, Biomaterials Res., 24, 5 (2020)

[5] Choi JS, Oh SH, Kim YM, Lim JY, Tissue Eng. Regen. Med., 17, 651 (2020)

[6] Min SJ, Kim SW, Lee TH, Kwon IK, Son TI, Polym. Korea, 44(4), 451 (2020)

[7] Sung YK, Kim SW, Biomaterials Res., 24, 1 (2020)

[8] Yang JM, Kim HJ, Cho BK, Polym. Korea, 44, 21 (2020)

[9] Hong BM, Park SA, Park WH, Biomaterials Res., 23, 1 (2019)

[10] Xu X, Jha AK, Harrington DA, Farach-Carson MC, Jia X, Soft Mater., 8, 3280 (2012)

[11] Khunmanee S, Jeong Y, Park H, J. Tissue Eng., 8, 2041731417726464 2017.

[12] Yang B, Guo X, Zang H, Liu J, Carbohydr. Polym., 131, 233 (2015)

[13] Shimojo AA, Pires A, Lichy R, Santana MH, J. Brazil. Chem. Soc, 26, 506 (2015)

[14] Ibrahim S, Kang QK, Ramamurthi A, J. Biomed. Mater. Res. A, 94, 355 (2010)

[15] Calles JA, Ressia JA, Llabot JM, Valles EM, Palma SD, Sci. Pharm., 84, 61 (2016)

[16] Kirk JF, Ritter G, Finger I, Sankar D, Reddy JD, Talton JD, et al>, Biomatter, 3, e25633 (2013)

[17] Cao W, Sui J, Ma M, Xu Y, Lin W, Chen Y, Man Y, Sun Y, Fan Y, Zhang X, J. Mater. Chem. B, 7, 4413 (2019)

[18] Nakajima N, Ikada Y, Bioconjugate Chem., 6, 123 (1995)

[19] Bulpitt P, Aeschlimann D, J. Biomed. Mater. Res. A, 47, 152 (1999)

[20] Kunishima M, Kawachi C, Monta J, Terao K, Iwasaki F, Tani S, Tetrahedron, 55, 13159 (1999)

[21] Farkas P, Bystricky S, Carbohydr. Polym., 68, 187 (2007)

[22] Raw SA, Tetrahedron Lett., 50, 946 (2009)

[23] D’este M, Eglin D, Alini M, Carbohydr. Polym., 108, 239 (2014)

[24] Til H, Falke H, Prinsen M, Willems M, Food Chem. Toxicol., 35, 337 (1997)

[25] Farriol M, Segovia-Silvestre T, Castellanos JM, Venereo Y, Orta X, Nutrition, 17, 934 (2001)

[26] Seliktar D, Science, 336(6085), 1124 (2012)

[27] Tomihata K, Ikada Y, J. Biomed. Mater. Res. B, 37, 243 (1997)