Chitosan-Catechin Conjugates의 제조 및 이들의 항산화 활성

제재영 ( Jae Young Je ) 한국키틴키토산학회 2012 한국키틴키토산학회지 Vol.17 No.3

본 연구에서는 천연고분자인 키토산에 카테킨을 도입하여 새로운 형태의 항산화 활성이 우수한 chitosan-catechin conjugates를 제조하였다. Chitosan-catechin conjugates의 확인은 1H NMR 및 DSC를 이용하였으며, 이들의 항산화 활성을 in vitro에서 검토하였다. 제조한 chitosan-catechin conjugates의 카테킨 함량은 몰 비율이 높을수록 높게 나왔으나, 경제적인 측면을 고려할 경우 몰 비율이 1:0.1이 적절한 것으로 판단되었다. DPPH, H2O2 및 ABTS+라디칼 소거능 및 환원력을 측정한 결과 카테킨의 함량이 가장 높은 conjugate에서 가장 우수한 항산화 활성을 나타났으며, 카테킨이 키토산에 도입됨으로써 항산화 활성이 향상됨을 알 수 있었다. 또한 지질과산화 억제능에서도 우수한 활성을 나타내어 향후 건강기능식품소재 등으로 활용이 가능할 것으로 판단된다. In this study, chitosan-catechin conjugates were prepared by free radical-mediated grafting mechanism using different molar ratios of chitosan residue and catechin. The chitosan-catechin conjugates were verified using H NMR and differential scanning calorimetry, and catechin content in the chitosan-catechin conjugates was determined. Catechin content was increased with increasing molar ratio; however, grafting efficiency was decreased with increasing molar ratio. Antioxidant abilities of the chitosan-catechin conjugates were investigated using 2,2-diphenyl-1-picrylhydrazyl (DPPH), hydrogen peroxide, and 2-azinobis(3-ethylbenzthiazoline)-6-sulfonic acid (ABTS) cation radical scavenging assay, and reducing power. All the chitosancatechin conjugates showed increased antioxidant abilities compared to the unmodified chitosan (p < 0.05), and the chitosancatechin (III), which was prepared using 1:1 of molar ratio, showed the highest antioxidant activities. In addition, the chitosancatechin conjugates significantly (p < 0.05) inhibited the formation of malondialdehyde, which is an indicator of lipid peroxidation, in linoleic acid emulsion system.

Chitosan-phytochemical Conjugates의 항산화 및 항염증 활성

제재영 ( Jae Young Je ) 한국키틴키토산학회 2015 한국키틴키토산학회지 Vol.20 No.4

본 연구에서는 chitosan에 p-coumaric acid, quercetin 및 syringic acid를 도입하여 chitosan의 항산화 및 항염증 활성의 변화에 대하여 알아보았다. Quercetin과 syringic acid가 도입된 chitosan-quercetin과 chitosan-syringic acid에서는 이들이 도입되지 않은 plain chitosan보다 우수한 항산화 및 항염증 활성을 나타내었다. 따라서 이들 소재들을 식품첨가제 및 기능성 식품 소재로 활용 가능할 것으로 기대된다. Chitosan-phytochemical conjugates were prepared by free radical-mediated conjugation method. Successful preparation of chitosan-p-coumaric acid, chitosan-quercetin, and chitosan-syringic acid was confirmed using 1 H NMR spectroscopy and the contents of each p-coumaric acid, quercetin, and syringic acid in the conjugates were also determined. Three conjugates were subjected to evaluate antioxidant activity and inhibition activity for nitric oxide (NO) production in lipopolysaccharide-stimulated RAW264.7 macrophages. Chitosan-quercetin and chitosan-syringic acid exhibited significantly (p<0.05) enhanced antioxidant activities compared to plain chitosan, which is prepared without addition of phytochemicals, but chitosan-p-coumaric acid showed similar antioxidant activity than those of plain chitosan. In lipid peroxidation assay, chitosan-quercetin only showed significantly (p<0.05) enhanced lipid peroxidation inhibition activity. In inhibition activity for NO production, chitosan-quercetin and chitosan-syringic acid significantly (p<0.05) inhibited NO production in LPS-stimulated RAW264.7 macrophages.

Propionibacterium acnes에 의해 유도된 염증에 대한 Chitosan-phytochemical Conjugates의 항염증 효과

김지훈 ( Ji-hoon Kim ),제재영 ( Jae-young Je ),김영목 ( Young-mog Kim ) 한국수산과학회(구 한국수산학회) 2016 한국수산과학회지 Vol.49 No.5

Propionibacterium acnes infection in skin tissue often causes acne vulgaris, commonly characterized by inflammatory papules, pustules, and nodules. Chitosan and its derivatives possess strong anti-inflammatory effects. In this study, the anti-inflammatory activity of chitosan-phytochemical conjugates on P. acnes-infected human skin keratinocytes (HaCaT) was evaluated. We designed a model of P. acnes-induced inflammation in viable HaCaT cells. Nitric oxide (NO), an inflammatory marker, was successfully elevated by P. acnes infection in HaCaT cells in a dose-dependent manner. Furthermore, the levels of NO were reduced by treatment with chitosan-phytochemical conjugates (chitosan-caffeic acid, -ferulic acid and -sinapic acid) in a dose-dependent manner. Among these conjugates, chitosan-caffeic acid exhibited the strongest NO suppression in HaCaT cells infected with P. acnes. The results obtained in this study suggest that chitosan-phytochemical conjugates could be used as a potential therapeutic agent against acne vulgaris.

Collagenase로 유도한 가토의 슬관절염에서 TGF-β1과 키토산-TGF-β1 복합체의 영향

장의찬 ( Eui Chan Chang ),송광섭 ( Kwang Sup Song ),정호중 ( Ho Jung Jung ),박영욱 ( Young Uk Park ),김미경 ( Mi Kyung Kim ) 대한슬관절학회 2006 대한슬관절학회지 Vol.18 No.2

목적: Collagenase type II로 유도한 가토의 슬관절염에서 TGF-β1과 키토산-TGF-β1 복합체의 영향을 알아보고자 하였다. 대상 및 방법: 가토 15마리의 우측 슬관절에 Collagenase를 실험 1일 및 4일째 두 번 주입하여 관절염을 유발한 후 실험 10일째 TGF-β1 (15 ng/mL, 1 mL, 1군), 키토산-TGF-β1 복합체(Chitosan: 5.8 g/mL, TGF-β1: 15 ng/mL, 1 mL, 2군), PBS(1 mL, 3군)을 각각 5마리씩 같은 부위에 주입하여 1주 간격으로 슬관절의 염증 소견 및 절름거림의 정도를 관찰하였다. 실험 4주째 관절연골과 활액막 조직을 얻어 각 군간의 육안적(India ink), 조직학적 소견(H&E, Safranin-O)을 평가(Kikuchi 점수)하였다. 결과: 외양상 염증 소견은 collagenase를 주사 후 1주 후에 가장 심하였으며, 이후로 각 군간 의미있는 차이는 없었으며, 절름거림의 정도는 3군에서 심했다. 육안적 소견은 2군에서 우수하였으며, 이러한 차이는 조직학적 평가에서 더욱 뚜렷하였다(1군: 평균 14±2.7, 2군: 평균 10.3±1.2, 3군: 평균 20.3±3.7) (p<0.05). 결론: Collagenase로 유도한 가토의 슬관절염의 회복에 과정에서 TGF-β1 단독 사용보다 키토산-TGF-β1 복합체의 사용이 효과적인 것으로 생각된다. Purpose; To evaluate the effects of the chitosan-TGF-β1 conjugate and TGF-β1 on osteoarthritic rabbit cartilage induced by collagenase type II. Materials and Methods; Chemical arthritis was induced in right knee of 15 rabbits by injection of collagenase type II (4 mg/mL, 0.25 cc) twice at day 1 and 4, and then each 5 rabbits were treated by intra-articular injection of TGF-β1 (15 ng/mL, 1 mL, group I), chitosan-TGF-β1 conjugate (Chitosan: 5.8 g/mL, TGF-β1: 15 ng/mL, group II) and PBS (1 mL, group III) at 10 days later in the same knees. The appearance of the injected knee joints and lameness were observed weekly. The articular cartilage and synovium obtained at 4 weeks were analysed by gross findings (India ink), histological examinations (H&E, Safrainin O) and evaluation through the Kikuchi score. Result; The osteoarthritic sign about knee joints was the most severe at 1 weeks after the initiation of collagenase injections, but there wasere no difference of 3 groups in osteoarthritic sign and lameness was most severe in group 3. Group 2 showed the most superior results in gross findings using india ink and the these results were more prominent in the evaluation of histological scoring (average scores of each group I:14±2.7, II:10.3±1.2, III:20.3±3.7). Conclusion; These results might suggest that, in the rabbit osteoarthritic knee model induced by collagenase, Chitosan-TGF-β1 conjugate is more effective than only TGF-β1 in inhibiting cartilage degeneration.

Development of chitosan-catechol conjugates as mucoadhesive polymer: assessment of acute oral toxicity in mice

Loveleen Kaur,Ritu Raj,Ajay Kumar Thakur,Inderbir Singh 환경독성보건학회 2020 환경독성보건학회지 Vol.35 No.3

Development of modified polymers is the focused area of research for developing stable, effective, sustainable and economical polymeric materials for developing different drug delivery systems. Modification of chitosan by catechol functionalization is useful for developing chitosan derivative with the improved mucoadhesive property. Present study was designed to perform single dose acute oral toxicity on chitosan-catechol conjugates in Swiss albino mice as per international guidelines. Oral administration of modified chitosan did not exhibit any significant change in body weight, behavioural pattern, haematology, food intake and clinical symptoms in the experimental animals. In the histopathological study, no pathological changes were observed in the vital organs of mice when administered perorally with 300 mg/kg and 2,000 mg/kg body-weight doses of chitosan-catechol polymeric conjugates. Overall, it was concluded from the acute oral toxicity study that the oral administration of chitosan-catechol conjugates in mice did not produce any toxicity. Hence, chitosan-catechol conjugates could be designated and recommended as safe polymeric material for developing different drug delivery systems.

Cellular Radical Scavenging Activity of Chitosan-Catechin Conjugate in Mouse Macrophage Cells

( Young Sook Cho ),( Jae Young Je ) 한국키틴키토산학회 2013 한국키틴키토산학회지 Vol.18 No.3

Chitosan-catechin conjugate was prepared to examine cellular radical scavenging activity in mouse macrophage cells. No cytotoxicity was observed in the tested concentrations (50~400 μg/mL) of chitosan-catechin conjugate. Chitosan-catechin conjugate inhibited intracellular reactive oxygen species (ROS) formation determined by a fluorescence probe, 2,7-dichlorofluorescin diacetate (DCFH-DA), and the inhibition activity was 32.82% at 200 μg/mL, whereas unmodified chitosan showed 26.29% at the same concentration, indicating ROS scavenging activity was increased by conjugating catechin onto chitosan. Chitosan-catechin conjugate also inhibited lipid peroxidation by 1.07-fold compared with the control (non-treatment group). Furthermore, chitosancatechin conjugate up-regulated the expression of cellular glutathione level by 2.07-fold compared with non-treatment group.

Comparative study of photosensitizer loaded and conjugated glycol chitosan nanoparticles for cancer therapy

Lee, S.J.,Koo, H.,Jeong, H.,Huh, M.S.,Choi, Y.,Jeong, S.Y.,Byun, Y.,Choi, K.,Kim, K.,Kwon, I.C. Elsevier Science Publishers 2011 Journal of controlled release Vol.152 No.1

This study reports that tumor-targeting glycol chitosan nanoparticles with physically loaded and chemically conjugated photosensitizers can be used in photodynamic therapy (PDT). First, the hydrophobic photosensitizer, chlorin e6 (Ce6), was physically loaded onto the hydrophobically-modified glycol chitosan nanoparticles (HGC), which were prepared by self-assembling amphiphilic glycol chitosan-5β-cholanic acid conjugates under aqueous conditions. Second, the Ce6s were chemically conjugated to the glycol chitosan polymers, resulting in amphiphilic glycol chitosan-Ce6 conjugates that formed self-assembled nanoparticles in aqueous condition. Both Ce6-loaded glycol chitosan nanoparticles (HGC-Ce6) and Ce6-conjugated chitosan nanoparticles (GC-Ce6) had similar average diameters of 300 to 350nm, a similar in vitro singlet oxygen generation efficacy under buffer conditions, and a rapid cellular uptake profile in the cell culture system. However, compared to GC-Ce6, HGC-Ce6 showed a burst of drug release in vitro, whereby 65% of physically loaded drugs were rapidly released from the particles within 6.5h in the buffer condition. When injected through the tail vein into tumor bearing mice, HGC-Ce6 did not accumulate efficiently in tumor tissue, reflecting the burst in the release of the physically loaded drug, while GC-Ce6 showed a prolonged circulation profile and a more efficient tumor accumulation, which resulted in high therapeutic efficacy. These comparative studies with drug-loaded and drug-conjugated nanoparticles showed that the photosensitizer-conjugated glycol chitosan nanoparticles with excellent tumor targeting properties have potential for PDT in cancer treatment.

Effect of Bond Linkage on In vitro Drug Release and Anti-HIV Activity of Chitosan-Stavudine Conjugates

Rong Zeng,Renzhong Qiao,Zehu Wang,Hongran Wang,Liqiang Chen,Lin Yang,Liming Hu,Zelin Li 한국고분자학회 2012 Macromolecular Research Vol.20 No.4

Two kinds of chitosan-stavudine (d4T) conjugates, chitosan-O-isopropyl-5'-O-d4T monophosphate conjugate (Cs-P-d4T) with a phosphoramide linkage and chitosan-5'-O-succinyl-d4T conjugate (Cs-S-d4T) with a succinic spacer, were synthesized using an Atherton-Todd reaction and carbodiimide coupling reaction, respectively, and then structurally characterized. Their in vitro drug release behaviors and anti-human immunodeficiency virus (HIV) activity were investigated and compared. Both of the chitosan-d4T conjugates more strongly prefer to release corresponding d4T derivatives rather than free d4T in a prolonged manner but have different hydrolysis routes. The anti-HIV activity and cytotoxicity evaluated in the MT4 cell line revealed that the anti-HIV selectivity index was in the following order: Cs-P-d4T > d4T >> Cs-S-d4T since the released d4T-5'-(O-isopropyl) monophosphate from Cs-P-d4T can bypass the rate-limiting bottleneck of nucleoside phosphorylation, while the released 5'-O-succinyl-d4T from Cs-Sd4T has to be hydrolyzed to d4T and then successively phosphorylated to its active form to exert antiviral activity. The results suggested that constructing a chitosan-nucleoside reverse transcriptase inhibitor (NRTI) conjugate with a phosphoramide linkage may be an efficient approach for improving NRTI therapy efficacy in antiretroviral treatment.

Intra-articular delivery of kartogenin-conjugated chitosan nano/microparticles for cartilage regeneration

Kang, M.L.,Ko, J.Y.,Kim, J.E.,Im, G.I. IPC Science and Technology Press 2014 Biomaterials Vol.35 No.37

We developed an intra-articular (IA) drug delivery system to treat osteoarthritis (OA) that consisted of kartogenin conjugated chitosan (CHI-KGN). Kartogenin, which promotes the selective differentiation of mesenchymal stem cells (MSCs) into chondrocytes, was conjugated with low-molecular-weight chitosan (LMWCS) and medium-molecular-weight chitosan (MMWCS) by covalent coupling of kartogenin to each chitosan using an ethyl(dimethylaminopropyl) carbodiimide (EDC)/N-hydroxysuccinimide (NHS) catalyst. Nanoparticles (NPs, 150 +/- 39 nm) or microparticles (MPs, 1.8 +/- 0.54 μm) were fabricated from kartogenin conjugated-LMWCS and -MMWCS, respectively, by an ionic gelation using tripolyphosphate (TPP). The in vitro release profiles of kartogenin from the particles showed sustained release for 7 weeks. When the effects of the CHI-KGN NPs or CHI-KGN MPs were evaluated on the in vitro chondrogenic differentiation of human bone marrow MSCs (hBMMSCs), the CHI-KGN NPs and CHI-KGN MPs induced higher expression of chondrogenic markers from cultured hBMMSCs than unconjugated kartogenin. In particular, hBMMSCs treated with CHI-KGN NPs exhibited more distinct chondrogenic properties in the long-term pellet cultures than those treated with CHI-KGN MPs. The in vivo therapeutic effects of CHI-KGN NPs or CHI-KGN MPs were investigated using a surgically-induced OA model in rats. The CHI-KGN MPs showed longer retention time in the knee joint than the CHI-KGN NPs after IA injection in OA rats. The rats treated with CHI-KGN NPs or CHI-KGN MPs by IA injection showed much less degenerative changes than untreated control or rats treated with unconjugated kartogenin. In conclusion, CHI-KGN NPs or CHI-KGN MPs can be useful polymer-drug conjugates as an IA drug delivery system to treat OA.

Controlled network structures of chitosan-poly(ethylene glycol) hydrogel microspheres and their impact on protein conjugation

Jung, Sukwon,Tang, Yi,Shim, Gyurak,Lee, Chang-Soo,Choi, Chang-Hyung,Yi, Hyunmin Elsevier 2018 Biochemical engineering journal Vol.135 No.-

<P><B>Abstract</B></P> <P>We demonstrate a facile approach to manufacture hydrogel microspheres with controlled and macroporous network structures via a simple yet versatile micromolding-based technique. Specifically, highly uniform poly(ethylene glycol) (PEG) hydrogel microspheres containing chemically functional chitosan are readily fabricated using the micromolding-based technique that utilizes surface tension-induced droplet formation of aqueous prepolymer solution followed by photo-induced interfacial polymerization. Network structures of the hydrogel microspheres are readily controlled by simple addition of inert porogen, which induces phase separation during the polymerization. Fluorescent labeling studies reveal tunable distribution of the chitosan within the PEG networks (i.e., uniform and core-shell like distributions) depending on the content of the porogen in the prepolymer solution. In addition, protein conjugation studies show tunable pore sizes and 3D network structures by adjusting content of the porogen, and formation of macroporous networks leading to significantly improved protein conjugation kinetics. The macroporous network structures are further supported with scanning electron microscopy (SEM) results that correspond well with confocal microscopy results. We believe that our fabrication strategy offers a simple and robust route to construct diverse 3D hydrogel network structures with chemical functionality, enabling production of a variety of biofunctionalized hydrogel microspheres that can be utilized in various biomedical applications.</P> <P><B>Highlights</B></P> <P> <UL> <LI> Hydrogel microspheres are fabricated by interfacial polymerization of static fluid. </LI> <LI> Hydrogel network structures are readily tuned by using inert porogen. </LI> <LI> Incorporated chitosan moieties in microspheres are used as a conjugation handle. </LI> <LI> Macroporous networks lead to substantially improved protein conjugation. </LI> </UL> </P> <P><B>Graphical abstract</B></P> <P>[DISPLAY OMISSION]</P>