Chitosan derivatives alter release profiles of model compounds from calcium phosphate implants

The aim of the current study was to evaluate the impact of chitosan derivatives, namely N-octyl-chitosan and N-octyl-O-sulfate chitosan, incorporated in calcium phosphate implants to the release profiles of model drugs. The rate and extent of calcein (on M.W. 650 Da) ED, and FITC-dextran (M.W. 40 kDa) on in vitro release were monitored by fluorescence spectroscopy. Results show that calcein release is affected by the type of chitosan derivative used. A higher percentage of model drug was released when the hydrophilic polymer N-octyl-sulfated chitosan was present in the tablets compared with the tablets containing the hydrophobic polymer N-octyl-chitosan. The release profiles of calcein or FD from tablets containing N-octyl-O-sulfate revealed a complete release for FD after 120 h compared with calcein where 20% of the drug was released over the same time period. These results suggest that the difference in the release profiles observed from the implants is dependent on the molecular weight of the model drugs. These data indicate the potential of chitosan derivatives in controlling the release profile of active compounds from calcium phosphate implants. (C) 2009 Elsevier Ltd. All rights reserved.

In vitro fermentation of chitosan derivatives by mixed cultures of human faecal bacteria

Stirred, pH controlled batch cultures were carried out with faecal inocula and various chitosans to investigate the fermentation of chitosan derivatives by the human gut flora. Changes in bacterial levels and short chain fatty acids were measured over time. Low, medium and high molecular weight chitosan caused a decrease in bacteroides, bifidobacteria, clostridia and lactobacilli. A similar pattern was seen with chitosan oligosaccharide (COS). Butyrate levels also decreased. A three-stage fermentation model of the human colon was used for investigation of the metabolism of COS. In a region representing the proximal colon, clostridia decreased while lactobacilli increased. In the region representing the transverse colon, bacteroides and clostridia increased. Distally a small increase in bacteroides occurred. Butyrate levels increased. Under the highly competitive conditions of the human colon, many members of the microflora, are unable to compete for chitosans of low, medium or high molecular weight. COS were more easily utilised and when added to an in vitro colonic model led to increased production of butyrate, but some populations of potentially detrimental bacteria also increased. (c) 2005 Elsevier Ltd. All rights reserved.

Thermal studies of chitin-chitosan derivatives

New poly(azo) amino-chitosan compounds were obtained from the azo coupling reaction of N-benzyl chitosan and diazonium salts. The thermal behavior of these compounds was studied by thermogravimetric analysis (TG), differential thermogravimetric analysis (DTG), TG coupled with a Fourier-transform infrared, and differential scanning calorimetry (DSC). TG/DTG curves of chitin-chitosan polymer showed two thermal events attributed to water loss and decomposition of the polysaccharide after cross-linking reactions. Thermal analysis of the poly(azo) amino-chitosan compounds showed that the decomposition temperatures decreased when compared to the starting chitin-chitosan and N-benzyl chitosan. DSC results showed an agreement with the TG/DTG analyses. Thermal behavior of poly(azo) amino-chitosans suggest that these compounds could be considered as potential thermal sensors. © 2012 Akadémiai Kiadó, Budapest, Hungary.

Relevance of molecular weight of chitosan and its derivatives and their antioxidant activities in vitro

The antioxidant potency of different molecular weight (DMW) chitosan and sulfated chitosan derivatives was investigated employing various established in vitro systems, such as superoxide (O-2(.-))/hydroxyl ((OH)-O-.) radicals scavenging, reducing power, iron ion chelating. As expected, we obtained several satisfying results, as follows: Firstly, low molecular weight chitosan had stronger scavenging effect on O-2(.-) and (OH)-O-. than high molecular weight chitosan. For example the O-2(.-) scavenging activity of low molecular weight chitosan (9 kDa) and high molecular weight chitosan (760 kDa) were 85.86 % and 35.50 % at 1.6 mg/mL, respectively. Secondly, comparing with DMW chitosan, DMW sulfated chitosans had the stronger inhibition effect on 0(2)(.-). At 0.05 mg/mL, the scavenging activity on O-2(.-) reached 86.26 %, for low molecular weight chitosan sulfate (9 kDa), but that of low molecular weight chitosan (9 kDa) was 85.86 % at 1.6 mg/mL. As concerning chitosan and sulfated chitosan of the same molecular weight, scavenging activities of sulfated chitosan on superoxide and hydroxyl radicals were more pronounced than that of chitosan. Thirdly, low molecular weight chitosan sulfate had more effective scavenging activity on 02 and (OH)-O-. than that of high molecular weight chitosan sulfate. Fourthly, DMW chitosans and sulfated chitosans were efficient in the reducing power, especially LCTS. Their orders were found to be LCTS > CTS4 > HCTS > CTS3 > CTS2 > CTS1 > CTS. Fifthly, CTS4 showed more considerable ferrous ion-chelating potency than others. Finally, the scavenging rate and reducing power of DMW chitosan and sulfated derivatives increased with their increasing concentration. Moreover, change of DMW sulfated chitosans was the most pronounced within the experimental concentration. However, chelating effect of DMW chitosans were not concentration dependent except for CTS4 and CTS1. (C) 2004 Elsevier Ltd. All rights reserved.

Novel materials based on chitosan, its derivatives and cellulose fibres

O presente trabalho tem como principal objectivo o desenvolvimento de novos materiais baseados em quitosano, seus derivados e celulose, na forma de nanofibras ou de papel. Em primeiro lugar procedeu-se à purificação das amostras comerciais de quitosano e à sua caracterização exaustiva em termos morfológicos e físicoquímicos. Devido a valores contraditórios encontrados na literatura relativamente à energia de superfície do quitosano, e tendo em conta a sua utilização como precursor de modificações químicas e a sua aplicação em misturas com outros materiais, realizou-se também um estudo sistemático da determinação da energia de superfície do quitosano, da quitina e seus respectivos homólogos monoméricos, por medição de ângulos de contacto Em todas as amostras comerciais destes polímeros identificaram-se impurezas não polares que estão associadas a erros na determinação da componente polar da energia de superfície. Após a remoção destas impurezas, o valor da energia total de superfície (gs), e em particular da sua componente polar, aumentou consideravelmente. Depois de purificadas e caracterizadas, algumas das amostras de quitosano foram então usadas na preparação de filmes nanocompósitos, nomeadamente dois quitosanos com diferentes graus de polimerização, correspondentes derivados solúveis em água (cloreto de N-(3-(N,N,N-trimetilamónio)-2- hidroxipropilo) de quitosano) e nanofibras de celulose como reforço (celulose nanofibrilada (NFC) e celulose bacteriana (BC). Estes filmes transparentes foram preparados através de um processo simples e com conotação ‘verde’ pela dispersão homogénea de diferentes teores de NFC (até 60%) e BC (até 40%) nas soluções de quitosano (1.5% w/v) seguida da evaporação do solvente. Os filmes obtidos foram depois caracterizados por diversas técnicas, tais como SEM, AFM, difracção de raio-X, TGA, DMA, ensaios de tracção e espectroscopia no visível. Estes filmes são altamente transparentes e apresentam melhores propriedades mecânicas e maior estabilidade térmica do que os correspondentes filmes sem reforço. Outra abordagem deste trabalho envolveu o revestimento de folhas de papel de E. globulus com quitosano e dois derivados, um derivado fluorescente e um derivado solúvel em água, numa máquina de revestimentos (‘máquina de colagem’) à escala piloto. Este estudo envolveu inicialmente a deposição de 1 a 5 camadas do derivado de quitosano fluorescente sobre as folhas de papel de forma a estudar a sua distribuição nas folhas em termos de espalhamento e penetração, através de medições de reflectância e luminescência. Os resultados mostraram que, por um lado, a distribuição do quitosano na superfície era homogénea e que, por outro lado, a sua penetração através dos poros do papel cessou após três deposições. Depois da terceira camada verificou-se a formação de um filme contínuo de quitosano sobre a superfície do papel. Estes resultados mostram que este derivado de quitosano fluorescente pode ser utilizado como marcador na optimização e compreensão de mecanismos de deposição de quitosano em papel e outros substratos. Depois de conhecida a distribuição do quitosano nas folhas de papel, estudou-se o efeito do revestimento de quitosano e do seu derivado solúvel em água nas propriedades finais do papel. As propriedades morfológicas, mecânicas, superficiais, ópticas, assim como a permeabilidade ao ar e ao vapor de água, a aptidão à impressão e o envelhecimento do papel, foram exaustivamente avaliadas. De uma forma geral, os revestimentos com quitosano e com o seu derivado solúvel em água tiveram um impacto positivo nas propriedades finais do papel, que se mostrou ser dependente do número de camadas depositadas. Os resultados também mostraram que os papéis revestidos com o derivado solúvel em água apresentaram melhores propriedades ópticas, aptidão à impressão e melhores resultados em relação ao envelhecimento do que os papéis revestidos com quitosano. Assim, o uso de derivados de quitosano solúveis em água em processos de revestimento de papel representa uma estratégia bastante interessante e sustentável para o desenvolvimento de novos materiais funcionais ou na melhoria das propriedades finais dos papéis. Por fim, tendo como objectivo valorizar os resíduos e fracções menos nobres da quitina e do quitosano provenientes da indústria transformadora, estes polímeros foram convertidos em polióis viscosos através de uma reacção simples de oxipropilação. Este processo tem também conotação "verde" uma vez que não requer solvente, não origina subprodutos e não exige nenhuma operação específica (separação, purificação, etc) para isolar o produto da reacção. As amostras de quitina e quitosano foram pré-activadas com KOH e depois modificadas com um excesso de óxido de propileno (PO) num reactor apropriado. Em todos os casos, o produto da reacção foi um líquido viscoso composto por quitina ou quitosano oxipropilados e homopolímero de PO. Estas duas fracções foram separadas e caracterizadas.

Synthesis, characterization and antifungal activity of quaternary derivatives of chitosan on Aspergillus flavus

Two series of new chitosan derivatives were synthesized by reaction of deacetylated chitosan (CH) with propyl (CH-Propyl) and pentyl (CH-Pentyl) trimethylammonium bromides to obtain derivatives with increasing degrees of substitution (DS). The derivatives were characterized by 1H NMR and potentiometric titration techniques and their antifungal activities on the mycelial growth of Aspergillus flavus were investigated in vitro. The antifungal activities increase with DS and the more substituted derivatives of both series, CH-Propyl and CH-Pentyl, exhibited antifungal activities respectively three and six times higher than those obtained with commercial and deacetylated chitosan. The minimum inhibitory concentrations (MIC) were evaluated at 24, 48 and 72h by varying the polymer concentration from 0.5 to 16g/L and the results showed that the quaternary derivatives inhibited the fungus growth at polymer concentrations four times lower than that obtained with deacetylated chitosan (CH). The chitosans modified with pentyltrimethylammonium bromide exhibited higher activity and results are discussed taking into account the degree of substitution (DS). © 2012 Elsevier GmbH.

Hydrophobic effect of amphiphilic derivatives of chitosan on the antifungal activity against aspergillus flavus and aspergillus parasiticus

Low molecular weight amphiphilic derivatives of chitosan were synthesized, characterized and their antifungal activities against Aspergillus flavus and Aspergillus parasiticus were tested. The derivatives were synthesized using as starting material a deacetylated chitosan sample in a two step process: the reaction with propyltrimethylammonium bromide (Pr), followed by reductive amination with dodecyl aldehyde. Aiming to evaluate the effect of the hydrophobic modification of the derivatives on the antifungal activity against the pathogens, the degree of substitution (DS1) by Pr groups was kept constant and the proportion of dodecyl (Dod) groups was varied from 7 to 29% (DS2). The derivatives were characterized by 1H-NMR and FTIR and their antifungal activities against the pathogens were tested by the radial growth of the colony and minimum inhibitory concentration (MIC) methods. The derivatives substituted with only Pr groups exhibited modest inhibition against A. flavus and A. parasiticus, like that obtained with deacetylated chitosan. Results revealed that the amphiphilic derivatives grafted with Dod groups exhibited increasing inhibition indexes, depending on polymer concentration and hydrophobic content. At 0.6 g/L, all amphiphilic derivatives having from 7.0 to 29% of Dod groups completely inhibited fungal growth and the MIC values were found to decrease from 4.0 g/L for deacetylated chitosan to 0.25-0.50 g/L for the derivatives. These new derivatives open up the possibility of new applications and avenues to develop effective biofungicides based on chitosan. © 2013 by the authors.

Antifungal Activity of Quaternized Carboxymethyl Chitosan

Quaternized carboxymethyl chitosan (QCMC) were synthesized and their antifungal activities against Alternaria Solani (A. Solani) and Physalospora piricola Nose (P. piricola Nose) were investigated. The results indicated that the quaternized carboxymethyl chitosan derivatives had better inhibitory effects than CMC, and the antifungal activities should be affected by the cation in these compounds.

The influence of molecular weight of quaternized chitosan on antifungal activity

Quaternized chitosan derivatives with different molecular weights were synthesized in the laboratory. Subsequent experiments were conducted to test their antifungal activities against Botrytis cinerea Pers. (B. cinerea pers.) and Colletotrichum lagenarium (Pass) Ell.et halst (C. lagenarium (Pass) EII.et halst). Our results indicate that quaternized chitosan derivatives have stronger antifungal activities than chitosan. Furthermore, quaternized chitosan derivatives with high molecular weight are shown to have even stronger antifungal activities than those with low molecular weight. (c) 2007 Published by Elsevier Ltd.

The influence of the cationic of quaternized chitosan on antifungal activity

Quatemized chitosan: N-(2-hydroxyl-phenyl)-NN-dimethyl chitosan (NHPDCS), N-(5-chloro-2-hydroxyl-phenyl)-NN-dimethyl chitosan (NCHPDCS), N-(2-hydroxyl-5-nitro-phenyl)-NN-dimethyl chitosan (NHNPDCS) and N-(5-bromic-2-hydroxyl-phenyl)-NN-dimethyI chitosan (NBHPDCS) were synthesized and their antifungal activities against Botrytis cinerea Pers. (B. cinerea Pers.) and Colletotrichum lagenarium (Pass) Ell.et halst (C. lagenarium (Pass) Ellet halst) were investigated. The results indicated that the quaternized chitosan derivatives had better inhibitory effects than chitosan, and the antifungal activities should be affected by the cation in these compounds. (C) 2007 Elsevier B.V. All rights reserved.

Relevance of molecular weight of chitosan-N-2-hydroxypropyl trimethyl ammonium chloride and their antioxidant activities

The antioxidant potency of high/low molecular weight quatemary chitosan derivatives was investigated employing various established systems in vitro, such as superoxide (O-2(center dot-)) and hydroxyl (center dot OH) radicals scavenging, reducing power and iron ion chelating. As expected, we obtained several satisfying results, as follow: firstly, low molecular weight quaternary chitosan had stronger scavenging effect on O-2(center dot-) and center dot OH than high molecular weight quaternary chitosan. Secondly, the reducing power of low molecular weight quaternary chitosan was more pronounced than that of high molecular weight quaternary chitosan. Thirdly, ferrous ion chelating potency were showed to increase first and decrease afterwards with increasing concentration for two kinds of quaternary chitosans, namely, they have not concentration-dependence. However, the scavenging rate and reducing power of high and low molecular weight quaternary chitosans increased with their increasing conc centrations, and hence were concentration-dependent. (c) 2007 Elsevier Masson SAS. All rights reserved.

Hydroxyl radicals scavenging activity of N-substituted chitosan and quaternized chitosan

N-substituted chitosan and quaternized chitosan were synthesized and their antioxidant activity against hydroxyl radicals was assessed, respectively. Compared with the antioxidant activity of chitosan, the results indicated that the two kinds of chitosan derivatives had different scavenging ability on hydroxyl radicals, which should be related to the form of amido in the two kinds of chitosan derivatives. (c) 2006 Elsevier Ltd. All rights reserved.

Synthesis and evaluation of diethylethylamine-chitosan for gene delivery: Composition effects on the in vitro transfection efficiency

Chitosan has been indicated as a safe and promising polycation vector for gene delivery. However its low transfection efficiency has been a challenging obstacle for its application. To address this limitation, we synthesized chitosan derivatives which had increasing amounts of diethylethylamine groups (DEAE) attached to the chitosan main chain. The plasmid DNA VR1412 (pDNA), encoding the ß-galactosidase (ß-gal) reporter gene was used to prepare nanoparticles with the chitosan derivatives, and the transfection studies were performed with HeLa cells. By means of dynamic light scattering and zeta potential measurements, it was shown that diethylethylamine-chitosan derivatives (DEAEx-CH) were able to condense DNA into small particles having a surface charge depending on the polymer/DNA ratio (N/P ratio). Nanoparticles prepared with derivatives containing 15 and 25% of DEAE groups (DEAE15-CH and DEAE25-CH) exhibited transfection efficiencies ten times higher than that observed with deacetylated chitosan (CH). For derivatives with higher degrees of substitution (DS), transfection efficiency decreased. The most effective carriers showed low cytotoxicity and good transfection activities at low charge ratios (N/P). Vectors with low DS were easily degraded in the presence of lysozyme at physiological conditions in vitro and the nontoxicity displayed by these vectors opens up new opportunities in the design of DEAE-chitosan-based nanoparticles for gene delivery. © 2013 IOP Publishing Ltd.

壳聚糖衍生物抑菌性能及机理的研究

壳聚糖是一种天然的聚阳离子氨基多糖，由甲壳素经脱乙酰反应得到。作为天然可再生资源，壳聚糖以其特有的安全无毒、可生物降解、生物相容等特性，在农业、生物工程、制药、环保等领域引起广泛关注。随着对壳聚糖研究的不断深入，壳聚糖的抑菌活性及机理成为研究热点之一，但是，大多集中于壳聚糖分子量和脱乙酰度对其抑菌活性的影响，而对壳聚糖衍生物的抑菌性能及机理研究较少。本文合成了N–季铵盐，N,O–季铵盐，羧甲基壳聚糖希夫碱及其铜配合物，壳聚糖有机酸盐，并对它们的抑菌活性进行了研究，同时探讨了引入基团与抑菌活性之间的关系。 测定了壳聚糖及其季铵盐对黄瓜枯萎病菌、黄瓜炭疽病菌和桃褐腐病菌的抑制活性，结果表明壳聚糖季铵盐的抑制活性高于壳聚糖，其中，抑制活性最强的是N–异丁基–N,N–二甲基壳聚糖，当样品浓度为500μg/mL时，对三种病原菌的抑制率分别为：67.52%、70.42%和76.25%。壳聚糖分子带正电荷的氨基可以与菌体表面带负电的物质作用，导致菌体死亡，而壳聚糖季铵盐分子中的正电性可以加强这种作用，从而增强抑菌活性，而且随着正电性的增强，抑菌活性增强。 为了进一步研究壳聚糖季铵盐的正电性与其抑菌活性的关系，合成了含有氟、溴、氯吸电子基团苯环取代的壳聚糖季铵盐，通过计算分别得到取代基团的电负性以及氨基正电性。结果表明，随着取代基团电负性的增强，季铵盐氨基正电性增强，抑菌活性增强。其中，抑菌活性最强的是N–(2–氟)苄基–N,N–二甲基壳聚糖，当样品浓度为500μg/mL时，对三种病原菌的抑制率分别为：80.85%、100%和100%。 将壳聚糖的–NH2与有机酸的–H通过离子键结合制备得到了系列的固态壳聚糖有机酸盐：壳聚糖甲酸盐、壳聚糖乙酸盐、壳聚糖丙酸盐、壳聚糖丁酸盐和壳聚糖戊酸盐，抑菌活性结果表明，随着衍生物链长的增加，抑菌活性增强，而且衍生物抑菌活性与取代基团的电负性成正相关。 以羧甲基壳聚糖为原料，合成了羧甲基壳聚糖希夫碱及其铜配合物，各类衍生物的抑菌活性强弱顺序如下：羧甲基壳聚糖希夫碱铜配合物>羧甲基壳聚糖希夫碱>羧甲基壳聚糖，从而说明了基团的相互增效作用能增强衍生物的抑菌活性。 以N–季铵盐为原料，以环氧丙基三甲基氯化铵为改性剂合成了具有–N+(CH3)2R和–N+(CH3)3两种基团的N,O–季铵盐，抑菌活性结果表明，N,O–季铵盐的抑菌活性

三类新型壳聚糖衍生物的制备及其抑菌、抗氧化活性研究

本论文制备了三类新型壳聚糖衍生物，并对其抑菌及抗氧化活性进行了筛选与研究，具体结果如下。 通过在不同分子量的壳聚糖上引入1，3，5-噻二嗪硫酮首次合成壳聚糖噻二嗪硫酮衍生物（N-2-对氨基苯磺酰噻二嗪硫酮-壳聚糖（SATTCS）， N-2-邻甲基苯噻二嗪硫酮-壳聚糖（TITTCS）, N-2-邻氯苯噻二嗪硫酮-壳聚糖（CITTCS））；通过将羟丙基壳聚糖与不同的醛反应首次合成了羟丙基壳聚糖希夫碱（羟丙基壳聚糖水杨醛希夫碱（HHPCS）, 羟丙基壳聚糖5-氯水杨醛希夫碱（HCHPCS）, 羟丙基壳聚糖5-硝基水杨醛希夫碱（HNHPCS）, 羟丙基壳聚糖5-溴水杨醛希夫碱（HBHPCS）, 羟丙基壳聚糖糠醛希夫碱（FHPCS））；通过将不同分子量的壳聚糖与乙酰芳香胺反应首次合成了壳聚糖乙酰芳香胺衍生物（壳聚糖乙酰苯胺（ABCS）, 壳聚糖乙酰芳邻氯苯胺（ACCS），壳聚糖乙酰芳对甲苯胺（TDCS）, 壳聚糖乙酰甲萘胺（MDACS））。采用红外、元素分析等方法进行了结构鉴定。 通过对金黄色葡萄球菌、八叠菌、大肠杆菌、绿脓杆菌，芦笋茎枯病菌等一些真菌的抑菌活性研究表明：壳聚糖噻二嗪硫酮衍生物、壳聚糖乙酰芳香胺衍生物具有较好的抑制细菌的活性，且衍生物对金黄色葡萄球菌的抑制效果最好。分子量不同，抑菌活性也不同。其中，SATTCS和 LMANCS对细菌的抑制效果最好， SATTCS和TDCS的对真菌的抑菌活性最为显著。 抗氧化活性筛选结果表明：壳聚糖噻二嗪硫酮衍生物和羟丙基壳聚糖希夫碱具有很强的清除羟自由基的能力，清除能力全部优于Vc: 壳聚糖噻二嗪硫酮衍生物还具有较强的还原能力,其中TITTCS的还原能力比Vc强；并且，低分子量的壳聚糖噻二嗪硫酮衍生物的清除羟自由基的能力以及还原能力略强于高分子量的壳聚糖噻二嗪硫酮衍生物。 通过本研究工作制备了三类新型壳聚糖衍生物；生物活性研究表明：壳聚糖噻二嗪硫酮衍生物和壳聚糖乙酰芳香胺衍生物在抑菌方面具有应用潜力。研究成果对深入开发利用甲壳质资源，促进我国海洋药物的发展具有重要的意义。