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Chitooligosaccharide (COS) is a derivative of chitosan, which is a natural macromolecular compound. COS has been shown effects in an inflammatory response. Recent reports show that COS derivatives have enhanced anti-inflammatory activity by inhibiting intracellular signals. Evaluation of the anti-inflammatory effect of caffeic acid conjugated COS chain (CA-COS) was performed in this study. The effects of CA-COS on the inflammatory response were demonstrated in lipopolysaccharide-stimulated RAW264.7 macrophages. The results showed that CA-COS inhibited nitric oxide (NO) production and downregulated the gene expression of nitric oxide synthase (iNOS), and cytokines such as tumor necrosis factor-alpha (TNF-α), IL-1β, and IL-6 without cytotoxic effect. In addition, western blot analysis showed that CA-COS inhibits the protein expression of iNOS and nuclear factor kappa B (NF-kB), including p50 and p65, and mitogen-activated protein kinase (MAPK) signaling pathways. Collectively, these results provide clear evidence for the anti-inflammatory mechanism of CA-COS that show great potential as a novel agent for the prevention and therapy of inflammatory diseases., (© 2024 John Wiley & Sons Ltd.)

Various biological effects have been reported for sulfated chitosan oligosaccharides, but the molecular mechanisms of action of their anti-inflammatory effects are still unknown. This study aimed to evaluate the anti-inflammatory effects of sulfated chitosan oligosaccharides and to elucidate the possible mechanisms of action. The results showed that pretreated low molecular weight sulfated chitosan oligosaccharides inhibited the production of nitric oxide (NO) and inflammatory cytokines such as IL-6 and TNF-α in lipopolysaccharide (LPS)-activated RAW264.7 cells. The sulfated chitosan oligosaccharides also suppressed inducible nitric oxide synthase (iNOS), phosphorylation of JNK and translocation of p65, a subunit of NF-κB, into the nucleus by inhibiting degradation of IκB-α. Our investigation suggests sulfated chitosan oligosaccharides inhibit IL-6/TNF-α in LPS-induced macrophages, regulated by mitogen-activated protein kinases (MAPKs) pathways dependent on NF-κB activation.

Chitin is a natural polysaccharide of major importance. This biopolymer is synthesized by an enormous number of living organisms; considering the amount of chitin produced annually in the world, it is the most abundant polymer after cellulose. The most important derivative of chitin is chitosan, obtained by partial deacetylation of chitin under alkaline conditions or by enzymatic hydrolysis. Chitin and chitosan are known to have important functional activities but poor solubility makes them difficult to use in food and biomedicinal applications. Chitooligosaccharides (COS) are the degraded products of chitosan or chitin prepared by enzymatic or chemical hydrolysis of chitosan. The greater solubility and low viscosity of COS have attracted the interest of many researchers to utilize COS and their derivatives for various biomedical applications. In light of the recent interest in the biomedical applications of chitin, chitosan, and their derivatives, this review focuses on the preparation and biological activities of chitin, chitosan, COS, and their derivatives.

Eight kinds of phenolic acid conjugated chitooligosaccharides (COSs) were synthesized using hydroxyl benzoic acid and hydroxyl cinnamic acid. These phenolic acid conjugated-COSs with different substitution groups, including p-hydroxyl, 3,4-dihydroxyl, 3-methoxyl-4-hydroxyl and 3,5-dimethoxyl-4-hydroxy groups, were evaluated for their inhibitory activities against β-site amyloid precursor protein (APP)-cleaving enzyme (BACE) and inhibited BACE with a ratio of 50.8%, 74.8%, 62.1%, 64.8% and 42.6%, respectively at the concentration of 1,000 μg/mL. BACE is a critical component to reduce the levels of Aβ amyloid peptide in Alzheimer's disease (AD) which is based on the amyloid cascade theory in the brain, as this protease initiates the first step in Aβ production. Among them, Caffeic acid conjugated-COS (CFA-COS) was further analysed to determine mode of inhibition of BACE and it showed non-competitive inhibition. Hence in this study, we suggest that CFA-COS derivatives have potential to be used as novel BACE inhibitors to reduce the risk of AD.

Background/purpose: In the present study, the effect of 3-5 kDa chitooligosaccharide (COS) on homeostasis between the expression of collagen-degrading matrix metalloproteinases (MMPs) and collagen synthesis was investigated using ultraviolet (UV)-A irradiated dermal fibroblasts., Methods: UV protection imparted by 3-5 kDa COS was measured by examining the UV absorption spectrum. Collagenase MMP secretion was examined using an enzyme-linked immunosorbent assay. The levels of collagenases and collagen synthetic markers were determined by employing the reverse transcriptase-polymerase chain reaction and Western blot analysis., Results: The 3-5 kDa COS not only absorbed UV-A and UV-B light but also inhibited collagenase (MMP-1, MMP-8, and MMP-13) and gelatinase (MMP-2 and MMP-9) MMP expression. The suppression of MMP expression was found to be due to an increase in expression of the tissue inhibitors of MMP (TIMP)-1 and TIMP-2. Treatment with 3-5 kDa COS enhanced collagen synthetic markers such as procollagen, type I, III, and IV collagens in UV-A-irradiated dermal fibroblasts. Furthermore, the effects of 3-5 kDa COS on collagen degradation and collagen synthesis in UV-A irradiated dermal fibroblasts were regulated via the inhibition of activating protein-1 (AP-1) signaling., Conclusion: Our results suggest that 3-5 kDa COS can be used to develop as topical applications for antiphotoaging cosmeceuticals as it enhances collagen synthesis., (© 2012 John Wiley & Sons A/S.)

In this study we propose a novel anticancer agent using hetero-chitooligosaccharide (hetero-COS). To examine the possibility of the hetero-COS as a anticancer agent, we prepared nine kinds of hetero-COS with relatively higher molecular weights (90, 75 and 50-COS I, 5-10kDa), medium molecular weights (90, 75 and 50-COS II, 1-5kDa), and lower molecular weights (90, 75 and 50-III, below 1kDa), and their anticancer properties were investigated on HL-60 cells using flow cytometry and morphological analysis. The results obtained indicate that 90-COS III, which is relatively higher degree of deacetylation and lower molecular weights, showed the highest anticancer activity, and the data showed the anticancer property of the hetero-COSs depended on their degree of deacetylation values and molecular weight., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

In this study, a bioactive derivative of chitooligosaccharides (3-5 kDa) was synthesized via grafting of gallic acid onto chitooligosaccharides (G-COS) to enhance anti-allergic activity. Hence, G-COS was evaluated for its capabilities against allergic reactions in RBL-2H3 mast cells sensitized with dinitrophenyl-specific immunoglobulin E antibody and stimulated by antigen dinitrophenyl-bovine serum albumin. It was revealed that G-COS exhibited significant inhibition on histamine release and production as well as intracellular Ca(2+) elevation at the concentration of 200μg/ml. Likewise, the suppressive effects of G-COS on expression and production of interleukin (IL)-4 and tumor necrosis factor (TNF)-α were evidenced. Moreover, G-COS treatment caused a remarkable blockade on degradation of inhibitory κB-α (IκB-α) protein, translocation of nuclear factor (NF)-κB, and phosphorylation of mitogen-activated protein kinases (MAPKs). Notably, the inhibitory activities of G-COS on allergic reactions were found as a consequence of suppression of FcεRI expression in antigen-stimulated cells. Accordingly, G-COS was suggested to be a promising candidate of novel inhibitors against allergic reactions., (Copyright © 2012 Elsevier B.V. All rights reserved.)

In this study, eight kinds of phenolic acid conjugated chitooligosaccharides (PA-c-COSs) with different substitution groups, including p-hydroxyl {hydroxybenzoic acid-c-COS (HBA-c-COS), p-coumaric acid-c-COS (PCA-c-COS)}, 3,4-dihydroxyl {protocatechuic acid-c-COS (PTA-c-COS), caffeic acid-c-COS (CFA-c-COS)}, 3-methoxyl-4-hydroxyl {vanillic acid-c-COS (VNA-c-COS), ferulic acid-c-COS (FRA-c-COS)} and 3,5-dimethoxyl-4-hydroxy {syringic acid-c-COS (SRA-c-COS), sinapinic acid-c-COS (SNA-c-COS)}, were prepared by amide coupling reaction. Their antioxidant properties were evaluated using several in vitro models such as 2,2-diphenyl-1-picrylhydrazyl (DPPH), hydroxyl (OH) and nitric oxide (NO) radicals scavenging and reducing power assays. The structures of the synthesized compounds were confirmed by UV, FT-IR and (1)H NMR data. CFA-c-COS showed 81.6% and 89.8% scavenging against DPPH and NO radical formation, respectively. CFA-c-COS also showed higher reducing power and hydroxyl radical scavenging activity compared to those of other compounds. Hence, CFA-c-COS can be a potential antioxidant compound., (Crown Copyright © 2012. Published by Elsevier B.V. All rights reserved.)

The aim of this study is to investigate the inhibitory effects of aminoethyl-chitooligosaccharides (AE-COS) on oxidative stress in mouse macrophages (RAW 264.7 cells). The inhibitory effects of AE-COS on DNA and protein oxidation were studied in RAW 264.7 cells. Furthermore, free radical scavenging effect of AE-COS were determined in RAW264.7 cells by 2',7'-dichlorofluorescein (DCF) intensity and intracellular glutathione (GSH) level. AE-COS also inhibited myeloperoxidase (MPO) activity in human myeloid cells (HL-60). These results suggest that AE-COS acts as a potential free radical scavenger in RAW 264.7 cells., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Chitooligosaccharides (COS), the hydrolyzed product of chitosan and its derivatives, are known to have interesting pharmaceutical and medicinal applications due to its high solubility, non-toxicity, and increased functionality. Among them sulfated chitooligosaccharides (SCOSs) have been identified to possess enhanced biological activities. This study reports the effects of SCOSs with different molecular weights on the degradation of articular cartilage through unregulated collagenase expression. The results indicated that the SCOS II (3-5kDa) effectively inhibited the expressions of collagenases 1 and 3 and thereby prevented TNF-α induced degradation of collagen in human chondrosarcoma cells (SW-1353). Moreover, the signaling cascade responsible for this effect was found as SCOS II mediated suppression of NF-κB activation. Based on these data, it can be concluded that SCOS II prevented collagen degradation by inhibiting collagenases 1 and 3 via suppressing TNF-α induced NF-κB signaling. We suggest that SCOS II can be further studied as a potential candidate for the treatment of arthritis., (Crown Copyright © 2012. Published by Elsevier Ltd. All rights reserved.)

Chitooligosaccharide (COS), a derivative of chitosan, can be produced by either chemical or enzymatic hydrolysis. Looking back through past research, several technological approaches have been taken to prepare COSs, and enzymatic approaches are favorable due to their environmentally friendly methods, safety, and a lack of toxicity. Similar to chitosan, COSs can also be considered potential nutraceuticals due to their versatile biological activities, water-soluble properties, and absorption properties in the intestine. Therefore, this chapter provides certain methods for the preparation of COSs and addresses their biological properties such as antihypertensive, antioxidant, antitumor, antimicrobial, and other select biological activities., (Copyright © 2012 Elsevier Inc. All rights reserved.)

As a continuation of our research on chitooligosaccharides (COS), this study focused on the protective effect of COS of various molecular weights (1-3, 3-5, and 5-10 kDa) on cellular damage caused by ultraviolet B (UVB)-induced oxidative stress in human dermal fibroblast cells. The results show that the protective effect of COS on UVB-stressed human fibroblasts was dependent on molecular weight. COS suppressed UVB irradiation-induced reactive oxygen species generation and DNA damage, accompanied by the downregulation of matrix metalloproteinase (MMP)-1 and MMP-13. In a comparative analysis, COS (3-5 kDa) exhibited the most potent protective effect on UVB-stressed fibroblasts. The presence of COS (3-5 kDa) attenuated UVB-induced collagenolytic MMP production and collagen degradation. The photoprotective activity of COS (3-5 kDa) was confirmed by transcriptional phosphorylation of mitogen-activated protein kinase-responsive signaling pathways.

Chitooligosaccharides (COS), depolymerized products of chitosan, has received considerable attention as bioactive material due to their biocompatible, biodegradable, non-toxic and non-allergenic natures. In this study, COS of four different molecular weight ranges (<1, 1-3, 3-5 and 5-10 kDa) were investigated for their abilities to modulate inflammatory mediators in lipopolysaccharides (LPS)-stimulated BV2 microglia. At the concentration of 500 μg/ml, COS attenuate the productions of nitric oxide (NO) and prostaglandin E(2) (PGE(2)) by inhibiting inducible NO synthase (iNOS) and cyclooxygenase-2 (COX-2) expressions. Furthermore, the release and expression levels of inflammatory cytokines; including tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) and interleukin-1β (IL-1β) were also attenuated by COS. Notably, the inhibitory activity of COS depends significantly on its molecular weight, with lower molecular weight showed higher activity. In addition, the suppressive effects on the phosphorylation of JNK and p38 mitogen-activated protein kinase (MAPK) by COS were confirmed. These results indicate that COS could be used as an inhibitor in regulating microglial inflammatory responses. Moreover, COS may assist therapeutic treatment of neurodegenerative diseases which accompanied with microglial activation., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Chitooligosaccharides are nontoxic and water-soluble compounds obtained by enzymatic degradation of chitosan, which is derived from chitin by a deacetylation process. Chitooligosaccharides possess broad range of activities such as antitumour, antifungal, antibacterial activities. Sulfated chitooligosaccharides (SCOSs) with different molecular weights were synthesized by a random sulfation reaction. In the present study, anti-HIV-1 properties of SCOSs and the impact of molecular weight on their inhibitory activity were investigated. SCOS III (MW 3-5 kDa) was found to be the most effective compound to inhibit HIV-1 replication. At nontoxic concentrations, SCOS III exhibited remarkable inhibitory activities on HIV-1-induced syncytia formation (EC(50) 2.19 microg/ml), lytic effect (EC(50) 1.43 microg/ml), and p24 antigen production (EC(50) 4.33 microg/ml and 7.76 microg/ml for HIV-1(RF) and HIV-1(Ba-L), respectively). In contrast, unsulfated chitooligosaccharides showed no activity against HIV-1. Furthermore, it was found that SCOS III blocked viral entry and virus-cell fusion probably via disrupting the binding of HIV-1 gp120 to CD4 cell surface receptor. These results suggest that sulfated chitooligosaccharides represent novel candidates for the development of anti-HIV-1 agent., (Copyright 2009 Elsevier Ltd. All rights reserved.)

Clinical applications of acetylcholinesterase (AChE) inhibitors are widespread in Alzheimer's sufferers in order to activate central cholinergic system and alleviate cognitive deficits by inhibiting the hydrolysis of acetylcholine. In this study, six kinds of chitooligosaccharides (COSs) with different molecular weight and degree of deacetylation were examined for their inhibitory effects against AChE. The 90-COSs exhibited potent AChE inhibitory activities compared to 50-COSs, while 90-MMWCOS (1000-5000 Da) in the 90-COSs showed the highest activity. Cell culture experiment revealed that 90-MMWCOS suppressed the level of AChE protein expression and AChE activity induced by Abeta(25-35) in PC12 cell lines.

Despite extensive study on biological activities of chitosan and chitooligosaccharides (COS), there is no experimental evidence available as to COS mediated inhibition of free radical damage in cellular oxidizing systems. In this study, radical scavenging efficacies of different molecular weight bearing COS were assessed and their intracellular radical scavenging effects were tested employing B16F1, murine melanoma cell line. The results exhibited appreciable suppression in occurrence of intracellular radical species in the presence of low molecular weight bearing COS (<1 kDa) confirming low molecular weight is important for observed activities in biological systems. However, DNA oxidation carried out in the presence of COS clearly exhibited that COS exert protective effect on oxidative damage of purified genomic DNA regardless of molecular weight. Low molecular weight bearing COS was observed to be successively participated in suppression of NF-kappaB gene promoter activity suggesting its capability to prevent oxidative stress related disease complications. Moreover, induction of intracellular glutathione (GSH) level in the presence of COS promoted the effectiveness of COS to act against cellular oxidative stress.

The objective of this study was to identify the cellular antioxidant effects of carboxylated chitooligosaccharides (CCOS), a chemically modified derivative of chitooligosaccharides (COS), by assessing oxidation inhibition potential on cellular biomolecules such as lipids, proteins, and direct scavenging of reactive oxygen species (ROS). Radical-mediated oxidation of cell membrane lipids and proteins was dose-dependently inhibited by CCOS, assessed by amount of lipid hydroperoxides and carbonyl carbon content in mouse macrophages, RAW264.7 cells. Further, CCOS inhibited myeloperoxidase (MPO) activity in human myeloid cells (HL60) suggesting indirect possibility of inhibiting generation of reactive oxygen species (ROS) such as superoxide radicals, H(2)O(2) and HOCl. Direct radical scavenging studies carried out with DCFH-DA fluorescence probe concluded that CCOS can act as a potent radical scavenger in cells.

Matrix metalloproteinases (MMPs) play a fundamental role in invasion and metastasis of tumor and, recent advances in medicinal chemistry have approached designing of MMP inhibitors with desired structural properties, selectivity and bioavailability. In the present study, novel low-molecular-weight carboxylated chitooligosaccharides (CCOS) were evaluated for their MMP-9 inhibitory effect on human fibrosarcoma cell line (HT1080). In zymography experiments, a clear dose-dependent inhibition on MMP-9 mediated gelatinolytic activities were observed in HT1080 cells following treatment with CCOS. Further, transfection studies carried out with MMP-9 and AP-1 reporter constructs suggested that the observed reduction in MMP-9 expression was due to down-regulation of MMP-9 transcription that mediated via inhibition of AP-1. Moreover, expression of c-Fos protein levels in cytoplasm and nucleus confirmed that CCOS could inhibit AP-1 expression but not its translocation. However, in the presence of CCOS, NF-kappaB and TIMP-1 expression levels remained constant. More importantly, inhibition of MMP-9 expression clearly led to inhibit tumor invasiveness that was studied with reconstituted basement membrane matrix proteins coated synthetic membranes. Taken together, this study discusses MMP-9 inhibition potential of CCOS and their involvement to demote degradation and cellular invasion of extracellular matrix (ECM) and basement membrane. Thus, control of MMP-9 expression by CCOS has considerable significance for the regulation of tumor progression.

Matrix metalloproteinase-9 (MMP-9) has gelatinase activity and plays an important role in cancer invasion and metastasis. Therefore, inhibition of specific types of MMPs including MMP-9 has become an attractive target for therapeutic intervention. The aim of this study was to investigate the effect of chitooligosaccharides (COS) on activity and expression of MMP-9 in HT1080 cells. The inhibitory effect of COS with different molecular masses was examined by gelatin zymography, reverse transcriptase-polymerase chain reaction (RT-PCR), gene reporter assay, and Western blot analysis. MMP-9 inhibition in the presence of COS was clearly observed in gelatin zymography. Specifically, 1- to 3-kDa COS (COS-I) exhibited the highest inhibitory effect on MMP-9 activity in HT1080 cells among tested molecular mass fractions. It was also found that COS-I was capable of inhibiting both gene and protein expression of MMP-9 (P<0.01). These results suggest that low molecular mass COS can be considered as a potent inhibitor of MMP-9.

Even though several studies report the importance of chitosan derivatives for their anticancer activity, no clear information is available to describe the relationship between their charge properties and observed activities. In this research, differently charged chitooligosaccharide (COS) derivatives were synthesized and their anticancer activities were studied using three cancer cell lines, HeLa, Hep3B and SW480. Neutral red and MTT cell viability studies revealed that, highly charged COS derivatives could significantly reduce cancer cell viability, regardless to the positive or negative charge. Further, fluorescence microscopic observations and DNA fragmentation studies confirmed that the anticancer effect of these highly charged COS derivatives were due to necrosis. However, the exact molecular mechanism for anticancer activity of strongly charged COS compared to their poorly charged counterparts is not clear.

Chitosan polymer with deacetylation degree of 93% was hydrolyzed with an endo-type chitosanase (35,000 U/g protein) with substrate to enzyme ratio of 1 to 1.5 for 18 h in a batch reactor, and then the resultant hydrolysates were fractionated into four different molecular weights using an ultrafiltration (UF) membrane reactor system. An in vitro study elucidated that four kinds of chitooligosaccharides (COSs) could efficiently inhibit the formation of insoluble calcium salts in the neutral pH. In vivo effects of COSs on Ca bioavailability were further studied in the osteoporosis modeling rats induced by ovariectomy and concurrent low calcium intake. During the experimental period corresponding to the menopause with the osteoporosis disease, calcium retention was increased and bone turnover was decreased by COS IV supplementation in the ovariectomized (OVX) rats. After the low Ca diet, COS IV diet including both normal level of calcium and vitamin D significantly decreased calcium loss in feces and increased calcium retention compared to the control diet. The levels of femoral total calcium, bone mineral density (BMD), and femoral strength were also significantly increased by the COS IV diet in a similar level to those of CPP diet group. In the present study, the results proved the beneficial effects of low molecular chitooligosaccharide (COS IV) in preventing negative mineral balance.

In the present research, chitooligosaccharides (COS) were carboxylated with -COCH(2)CH(2)COO(-) groups to obtain specific structural features similar to Captopril. Angiotensin I converting enzyme (ACE) inhibitory activity of carboxylated COS was studied and observed to enhance its activity with increased substitution degree. Further, Lineweaver-Burk plot analysis revealed that inhibition was competitive via obligatory binding site of the enzyme. This was accompanied with substitution of positively charged quarternized amino groups to COS with different substitution degrees, in which negative impact on ACE inhibition was observed.

Nine kinds of hetero-chitooligosaccharides (hetero-COSs) with relatively higher molecular weights (90, 75 and 50-HMWCOS), medium molecular weights (90, 75 and 50-MMWCOS), and lower molecular weights (90, 75 and 50-LMWCOS) were prepared from partially deacetylated hetero-chitosans (90, 75 and 50% deacetylated chitosan), and their scavenging activities were investigated against 1,1-diphenyl-2-picrylhydrazyl (DPPH), hydroxyl, superoxide and carbon-centered radicals using electron spin resonance (ESR) spin-trapping technique. Superoxide, hydroxyl and carbon-centered radicals were generated from hypoxanthine-xanthine oxidase reaction, hydrogen peroxide-ferrous sulfate (Fenton reaction) and azo compound 2,2-azobis-(2-amidinopropane)-hydrochloride (AAPH), respectively. The ESR results revealed that 90-MMWCOS, which is having relatively medium molecular weights prepared from 90% deacetylated chitosan, showed the highest scavenging activity on all tested radicals. In addition, the radical scavenging activity of hetero-COSs increased with a dose-dependent manner, and it was dependent on their degree of deacetylation values and molecular weights.

Angiotensin I converting enzyme (ACE) inhibitory activity of hetero-chitooligosaccharides (hetero-COSs) prepared from partially different deacetylated chitosans was investigated. Partially deacetylated chitosans, 90, 75, and 50% deacetylated chitosan, were prepared from crab chitin by N-deacetylation with 40% sodium hydroxide solution for durations. In addition, nine kinds of hetero-COSs with relatively high molecular masses (5000-10 000 Da; 90-HMWCOSs, 75-HMWCOSs, and 50-HMWCOSs), medium molecular masses (1000-5000 Da; 90-MMWCOSs, 75-MMWCOSs, and 50-MMWCOSs), and low molecular masses (below 1000 Da; 90-LMWCOSs, 75-LMWCOSs, and 50-LMWCOSs) were prepared using an ultrafiltration membrane bioreactor system. ACE inhibitory activity of hetero-COSs was dependent on the degree of deacetylation of chitosans. 50-MMWCOSs that are COSs hydrolyzed from 50% deacetylated chitosan, the relatively lowest degree of deacetylation, exhibited the highest ACE inhibitory activity, and the IC(50) value was 1.22 +/- 0.13 mg/mL. In addition, the ACE inhibition pattern of the 50-MMWCOSs was investigated by Lineweaver-Burk plots, and the inhibition pattern was found to be competitive.

The radical scavenging effects of chitooligosaccharides (COSs) on hydroxyl radical, superoxide radical, alkyl radical, and 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical were investigated using a spin-trapping electron spin resonance (ESR) method and compared with the ESR signal intensity. COSs exhibited strong scavenging activity on hydroxyl radical and superoxide radical and weak scavenging activity on alkyl radical and DPPH radical. The radical scavenging activity of COSs increased with increment of concentration, and it was also dependent on molecular weight. These results suggest that the scavenging activity of COSs is dependent on their molecular weights and tested radicals.

The fourth generation of bioethanol production is on a lookout for non-lignocellulosic biomass waste. One such candidate is chitin, the second most abundant biopolymer on earth. However, the crystalline nature of chitin hinders its application potential for bioethanol production. This limitation can be circumvented by hydrolysing this polymer into oligomers using chitinases. We used this hypothesis and isolated a Bacillus haynesii, a marine bacterium that utilizes colloidal chitin as a substrate and produces chitin oligosaccharides. Further, we utilized Mucor circinelloides to produce bioethanol using the chitin oligosaccharides in the shake flask. We investigated the effect of inoculum age, filling volume, different substrates, and substrate concentration on bioethanol production using Mucor circinelloides from Bacillus haynesii-produced chitin oligosaccharides. Bacillus haynesii demonstrated a maximum chitinase activity of 3.08 U/mL with specific activity of 96 U/mg at the 90th h. Chitin oligosaccharides produced by Bacillus haynesii were confirmed using mass spectrometry. Bioethanol concentration was determined using dichromate oxidation assay as well as gas chromatography. The research resulted in 7.4 g/L of ethanol from 30 g/L of chitin oligosaccharides, with a maximum ethanol yield of 0.25 g of ethanol/g substrate at the 55th h with 48 h inoculum in 80 mL of fermentation medium. Results suggest that chitin oligosaccharides from Bacillus haynesii are an effective and renewable substrate for bioethanol production. [ABSTRACT FROM AUTHOR]

Abstract: The purpose of this research is not only to produce two kinds of chitin oligosaccharides or N-acetyl chito-oligosaccharides (NA-COSs) with different molecular weights (MW) from crab chitin hydrolysis solution but also to determine their effect against oxidative stress in live cells. Two kinds of NA-COSs with MW 1–3kDa (NA-COS 1–3kDa) and below 1kDa (NA-COS<1kDa) were obtained using an ultrafiltration membrane system. They exhibited an inhibitory effect against DNA and protein oxidation. In addition, in their presence, intracellular glutathione (GSH) level and direct intracellular radical scavenging effect were significantly increased in a time-dependent manner in mouse macrophages (RAW 264.7) and rendered inhibitory effect against cellular oxidative stress. In particular, NA-COS 1–3kDa was more effective than NA-COS<1kDa in protein oxidation and production of intracellular free radicals in live cells. These results suggest that NA-COSs act as a potential scavenger against oxidative stress in cells. [Copyright &y& Elsevier]

Abstract: Chitooligosaccharides (COS) of three different molecular weight ranges (1–3kDa, 3–5kDa and 5–10kDa) were investigated for their abilities against allergic reactions in RBL-2H3 cells. At the high concentration of 1000μg/ml, COS with molecular weights of 1–3kDa greatly attenuated the histamine and β-hexosaminidase release and intracellular Ca2+ elevation induced by calcium ionophore A23187, at the rates of 32% and 34% release and 37% elevation, respectively. Furthermore, the inhibitory activities of COS on calcium ionophore A23187 plus phorbol 12-myristate 13-acetate (PMA)-induced production as well as mRNA expression of tumor necrosis factor (TNF)-α, interleukin (IL)-1β, IL-4, and IL-6 were evidenced. Notably, the suppressive effects on the phosphorylation of extracellular signal-regulated kinases (MEK/ERK) and p38 kinase by COS were confirmed. These results indicate that COS could be used as an inhibitor in regulating mast cell-mediated allergic inflammatory responses. [ABSTRACT FROM AUTHOR]

Abstract: In this study, three kinds of chitooligosaccharides (COS) with different substitution groups, including aminoethyl (AE), dimethylaminoethyl (DMEM) and diethylaminoethyl (DEAE) groups, were evaluated for their inhibitory activities against cholinesterase (acetylcholinesterase and butyrylcholinesterase). These COS derivatives were synthesized and their structures were characterized as AE-COS, DMAE-COS and DEAE-COS, respectively; the structures were elucidated via spectroscopic techniques (1H NMR). Of these, DMAE- and DEAE-COS were synthesized for the first time in this work. These COS derivatives evidenced potent acetylcholinesterase (AChE) inhibitory activities with IC50 values of 56.5±0.26, 24.1±0.39 and 9.2±0.33μg/ml, respectively; however, these compounds exhibited no activity against butyrylcholinesterase. These were further analyzed for their inhibitory pattern on AChE via Lineweaver–Burk plots. AE-COS showed non-competition type inhibition, and DMAE-COS and DEAE-COS exhibited competition type inhibition against AChE. In this study, we suggested that COS derivatives have potential as AChE inhibitors for preventing Alzheimer’s disease. [Copyright &y& Elsevier]

Abstract: The aim of this research is to identify the cellular antioxidant effects of chitin oligosaccharides (NA-COS; M w 229.21–593.12Da) produced by acidic hydrolysis of crab chitin. The inhibitory effect of NA-COS on myeloperoxidase (MPO) activity in human myeloid cells (HL-60) and oxidation of DNA and protein in mouse macrophages (Raw 264.7) were identified. Furthermore, their direct radical scavenging effect by 2′,7′-dichlorofluorescein (DCF) intensity and intracellular glutathione (GSH) level were significantly increased in a time dependent manner, respectively. These results suggest that NA-COS act as a potent antioxidant in live cells. [Copyright &y& Elsevier]

Abstract: Clinical applications of acetylcholinesterase (AChE) inhibitors are widespread in Alzheimer’s sufferers in order to activate central cholinergic system and alleviate cognitive deficits by inhibiting the hydrolysis of acetylcholine. In this study, six kinds of chitooligosaccharides (COSs) with different molecular weight and degree of deacetylation were examined for their inhibitory effects against AChE. The 90-COSs exhibited potent AChE inhibitory activities compared to 50-COSs, while 90-MMWCOS (1000–5000Da) in the 90-COSs showed the highest activity. Cell culture experiment revealed that 90-MMWCOS suppressed the level of AChE protein expression and AChE activity induced by Aβ25–35 in PC12 cell lines. [Copyright &y& Elsevier]

In this study, gallic acid-conjugated chitooligosaccharides (gallate-COS) was evaluated for its enhancing capabilities in free radical scavenging and anti-inflammatory activities in human lung epithelial A549 cells. It was found that gallate-COS possesses significantly high DPPH radical scavenging activity and protective effect against H 2 O 2 -induced DNA damage. Likewise, gallate-COS decreased the production of intracellular reactive oxygen species in H 2 O 2 -stimulated A549 cells. Moreover, the suppressive effects of gallate-COS on COX-2 expression and PGE 2 production from LPS-stimulated A549 cells were evidenced. Finally, the productions of cytokines including IL-8 and TNF-α were also inhibited by gallate-COS in a dose-dependent manner. Collectively, these results suggest that gallate-COS could be used as a functional ingredient with potent antioxidant and anti-inflammatory benefits. [ABSTRACT FROM AUTHOR]

Abstract: The aim of this study was to investigate the antioxidant and anti-inflammatory activities of aminoethyl-chitooligosaccharides (AE-COS) in murine microglial cells (BV-2). At the high concentration of 100μg/ml, the inhibition of reactive oxygen species, DNA, protein and lipid oxidation were determined in BV-2 cells, at the rates of 77%, 80%, 85% and 85%, respectively. Furthermore, AE-COS was studied for its capabilities against lipopolysaccharide-induced inflammatory responses in BV-2 cells. It was found that AE-COS reduced the level of nitric oxide (NO) and prostaglandin E2 production by diminishing the expression of inducible NO synthase and cyclooxygenase-2 without significant cytotoxicity. Moreover, the inhibitory activities of AE-COS on generation of tumor necrosis factor-alpha and interleukin-1beta were performed. Collectively, these results indicate that AE-COS possess potential antioxidant and anti-inflammatory activities in brain microglia, and could be a useful therapeutic agent for the treatment of neuroinflammatory diseases. [Copyright &y& Elsevier]

Abstract: The activation of mast cells by immunoglobulin E-mediated stimuli is considered as a central event in allergic responses. In this regard, chitosan oligosaccharides (COS) of two different molecular weight ranges (1–3kDa and 3–5kDa) were investigated for their capabilities against the activation of RBL-2H3 mast cell sensitized with dinitrophenyl-specific immunoglobulin E antibody and stimulated by antigen dinitrophenyl-bovine serum albumin. It was found that COS significantly inhibited RBL-2H3 cell degranulation via attenuating the releases of histamine and β-hexosaminidase. Moreover, the inhibitory activity of COS was accompanied by a reduction in intracellular Ca2+ elevation. Notably, the expression of immunoglobulin Fc epsilon receptor I (FcɛRI) in RBL-2H3 cells was down-regulated by COS treatment in a dose-dependent manner. The suppressive effect of COS on RBL-2H3 cell activation suggested that COS may be potential candidates of novel inhibitors against allergic reactions. [Copyright &y& Elsevier]

Abstract: This study explains the novel possibility of the involvement of gallic acid conjugated chitooligosaccharides (G-COS) in ameliorating cellular antioxidant activity. The free radicals scavenging activity of G-COS was assessed by using an electron spin resonance technique. In addition, the inhibition of reactive oxygen species, lipid, protein, and DNA oxidation were determined in human chondrosarcoma (SW1353 cells). Moreover, the antioxidative mechanism of G-COS was evaluated by protein and gene expression levels of antioxidant enzymes (superoxide dismutase and glutathione). Western blot analysis revealed that G-COS inhibit radical-mediated expression and activation of NF-κB proteins (transcription factor in the regulation of expression of numerous genes in the setting of the osteoarthritic response). Based on the results of this study, G-COS can be used as a scavenger in controlling free radicals that lead to damage to cellular systems in osteoarthritis. [Copyright &y& Elsevier]

Abstract: In this research, a novel derivative of chitooligosaccharides (COS) was synthesized by covalently linking gallic acid and COS (gallate-COS) via carbodiimide to improve cellular antioxidant activity. The direct intracellular radical scavenging effect of gallate-COS by 2′,7′-dichlorofluorescin diacetate (DCFH-DA) method, and the inhibition of oxidation of cellular macromolecules, such as DNA, protein and lipid in mouse macrophages (RAW264.7 cells) were determined. Furthermore, with the treatment of gallate-COS, the expression levels of antioxidant enzymes such as superoxide dismutase (SOD) and glutathione (GSH) were significantly increased according to reverse transcription polymerase chain reaction (RT-PCR) and Western blot analysis. In addition, gallate-COS decreased reactive oxygen species induced activation of the nuclear transcription factor (NF-κB). Collectively, these results suggest that gallate-COS can be used as a potential natural compound-based antioxidant in the functional food and pharmaceutical industries. [Copyright &y& Elsevier]

Abstract: In this study, the ability of aminoethylation of chitooligosaccharide (COS) to inhibit the proliferation of AGS human gastric adenocarcinoma cells was evaluated. Aminoderivatized COSs, aminoethyl-chitooligosaccharide (AE-COS), dimethyl aminoethyl-chitooligosaccharide (DMAE-COS) and diethyl aminoethyl-chitooligosaccharide (DEAE-COS), were synthesized and confirmed by their IR spectra results in comparison to previous study. Aminoderivatized chitooligosaccharides-induced cell death was characterized by cell viability assay, changes in nuclear morphology and changes in cell morphology. According to our results, all aminoderivatized COSs significantly induced cell death in AGS gastric cancer cells. Moreover, protein and gene expression levels of important regulators involved in apoptosis pathway such as Bcl-2, Bax, p53 and p21 were examined using RT-PCR and Western blot analysis. Aminoderivatized COSs showed dose- and time-dependent inhibition of AGS cancer cell proliferation. AE-COS and DEAE-COS showed the higher apoptotic activity than DMAE-COS. The present results suggest all three kinds of water-soluble aminoderivatized COSs have a promising potential as valuable as cancer chemopreventive agents. [ABSTRACT FROM AUTHOR]

Abstract: Chitooligosaccharides (COS) of four different molecular weight ranges (below 1kDa, 1–3kDa, 3–5kDa and 5–10kDa) were investigated for their ability to protect β-cells from damage induced by hydrogen peroxide. The viability of HIT-T15 pancreatic β-cells was found by MTT assay to be significantly reduced after 4h of treatment with 150μM H2O2. Chitooligosaccharides (500μg/ml) with molecular weights of 3–5kDa raised cell viability to 58.46% following H2O2 addition. The radical scavenging effect of chitooligosaccharides was examined by dichlorofluorescein assay. In addition to free radical scavenging, the effect of chitooligosaccharides on oxidative-stress related enzymes and genes was tested. In agreement with the expression levels of related genes, results of cell viability and cellular radical scavenging assays indicate that chitooligosaccharides protect β-cells from oxidative-stress-induced cellular damage, presumably due to radical scavenging activity. [Copyright &y& Elsevier]

Abstract: In this study, factors affecting anti-inflammatory effect of chitooligosaccharides (COSs) in lipopolysaccharides (LPS)-induced RAW264.7 macrophage cells were investigated. The inhibition of NO secretion by COSs revealed that 90-COSs (90% N-deacetylation) significantly inhibited NO secretion than those of 50-COSs (50% N-deacetylation), and 90-HMWCOS (5000–10,000Da) in the 90-COSs showed the highest inhibition activity. Furthermore, 90-HMWCOSs also found to inhibit LPS-stimulated production of PGE2, TNF-α and IL-6, as well as the expression of iNOS, COX-2, TNF-α, and IL-6. These results suggested that 90-HMWCOS may have anti-inflammatory effect via down-regulation of transcriptional and translational expression levels of TNF-α, IL-6 and iNOS and COX-2. [Copyright &y& Elsevier]