Multifaceted Carboxymethyl Chitosan Derivatives: Properties and Biomedical Applications [292] 9783031440991

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Advances in Polymer Science 292

R. Jayakumar Editor

Multifaceted Carboxymethyl Chitosan Derivatives: Properties and Biomedical Applications

Advances in Polymer Science Volume 292

Aims and Scope The series Advances in Polymer Science presents critical reviews of the present and future trends in polymer and biopolymer science. It covers all areas of research in polymer and biopolymer science including chemistry, physical chemistry, physics, and material science. The thematic volumes are addressed to scientists, whether at universities or in industry, who wish to keep abreast of the important advances in the covered topics. Advances in Polymer Science enjoys a longstanding tradition and good reputation in its community. Each volume is dedicated to a current topic, and each review critically surveys one aspect of that topic, to place it within the context of the volume. The volumes typically summarize the significant developments of the last 5 to 10 years and discuss them critically, presenting selected examples, explaining and illustrating the important principles, and bringing together many important references of primary literature. On that basis, future research directions in the area can be discussed. Advances in Polymer Science volumes thus are important references for every polymer scientist, as well as for other scientists interested in polymer science - as an introduction to a neighboring field, or as a compilation of detailed information for the specialist. Review articles for the individual volumes are invited by the volume editors. Single contributions can be specially commissioned. Readership: Polymer scientists, or scientists in related fields interested in polymer and biopolymer science, at universities or in industry, graduate students.

R. Jayakumar Editor

Multifaceted Carboxymethyl Chitosan Derivatives: Properties and Biomedical Applications With contributions by I. R. Antony
C. Arthi
E. T. Baran
T. Chaiwarit
P. Jantrawut
V. Jayachandran
R. Jayakumar
A. Jeyaseelan
K. Joy
G. Kasi
S. Li
F. G. L. Medeiros Borsagli
S. Meenakshi
S. S. Murugan
P. M. Nazreen
P. Panraksa
S. Parmaksız
M. Prabaharan
C. Rachtanapun
P. Rachtanapun
M. Nivedhitha Sundaram
C. Sairam Sundaram
D. Sathya Seeli
S. S¸enel
G. H. Seong
S. Thanakkasaranee
J. Tantala
N. Viswanathan
V. Venkatesan
R. Yu

Editor R. Jayakumar Centre for Nanosciences and Molecular Medicine, Amrita Institute of Medical Sciences and Research Centre Amrita Vishwa Vidyapeetham (University) Kochi, Kerala, India

ISSN 0065-3195 ISSN 1436-5030 (electronic) Advances in Polymer Science ISBN 978-3-031-44099-1 ISBN 978-3-031-44100-4 (eBook) https://doi.org/10.1007/978-3-031-44100-4 © The Editor(s) (if applicable) and The Author(s), under exclusive license to Springer Nature Switzerland AG 2024 This work is subject to copyright. All rights are solely and exclusively licensed by the Publisher, whether the whole or part of the material is concerned, specifically the rights of translation, reprinting, reuse of illustrations, recitation, broadcasting, reproduction on microfilms or in any other physical way, and transmission or information storage and retrieval, electronic adaptation, computer software, or by similar or dissimilar methodology now known or hereafter developed. The use of general descriptive names, registered names, trademarks, service marks, etc. in this publication does not imply, even in the absence of a specific statement, that such names are exempt from the relevant protective laws and regulations and therefore free for general use. The publisher, the authors, and the editors are safe to assume that the advice and information in this book are believed to be true and accurate at the date of publication. Neither the publisher nor the authors or the editors give a warranty, expressed or implied, with respect to the material contained herein or for any errors or omissions that may have been made. The publisher remains neutral with regard to jurisdictional claims in published maps and institutional affiliations. This Springer imprint is published by the registered company Springer Nature Switzerland AG The registered company address is: Gewerbestrasse 11, 6330 Cham, Switzerland Paper in this product is recyclable.

Preface

The volume titled “Multifaceted Carboxymethyl Chitosan Derivatives: Properties and Biomedical Applications” features the structure–property relationships and applications of these carboxymethyl chitosan derivatives in various fields. The individual chapters in this volume explain in detail about the properties, synthesis methods employed, interaction with metal ions. The antimicrobial activity of the CMC derivatives was covered in this volume. The chapters presented in this volume talk in detail about the development of functionalized carboxymethyl chitosan and its derivatives into various forms such as hydrogels, films, scaffolds, and composites. An important aspect, the application of these carboxymethyl chitosan derivatives in the pharmaceutical and cosmetics industry has also been highlighted. Certain chapters of this volume point out the extensive application of these derivatives used in a wide range of fields ranging from food packaging, in enhancing saltiness, as biosensors and also as wound dressing, etc. In addition to the aforementioned potential possibilities and applications, prospective current challenges in the development of biomedical products based on carboxymethyl chitosan and its derivatives have also been briefed. This volume will be advantageous to material/biomaterial scientists, biomedical engineers, chemists, and biotechnologists by providing a better insight on the subject, concerning the structure–property relationship, biological interaction, and applications of carboxymethyl chitosan and its derivatives in vivid fields. Kochi, India

R. Jayakumar

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Contents

Introductory Aspects of Carboxymethyl Chitosan Derivatives . . . . . . . . Irine Rose Antony and R. Jayakumar Preparation of Different Types of Carboxymethyl Chitosan Derivatives Irine Rose Antony and R. Jayakumar

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Carboxymethyl Chitosan Derivatives and Its Interaction with Metal Ions 31 Antonysamy Jeyaseelan, Natrayasamy Viswanathan, C. Sairam Sundaram, and S. Meenakshi Antimicrobial Properties of Carboxymethyl Chitosan Derivatives and Its Composites . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Vidyaalakshmi Venkatesan and R. Jayakumar

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Synthesis, Properties, and Applications of Carboxymethyl Chitosan-Based Hydrogels Rui Yu and Suming Li

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Carboxymethyl Chitosan Derivatives in Biosensing Applications . . . . . . Sesha S. Murugan, Gi Hun Seong, and Venkatesan Jayachandran

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Carboxymethyl Chitosan Derivatives in Blood Clotting . . . . . . . . . . . . . 109 C. Arthi, P. M. Nazreen, M. Nivedhitha Sundaram, and R. Jayakumar Electrospinning of Carboxymethyl Chitosan Derivatives-Based Nanofibers and Its Applications . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 125 C. Arthi and R. Jayakumar Carboxymethyl Chitosan-Based Materials in Packaging, Food, Pharmaceutical, and Cosmetics . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 139 P. Rachtanapun, C. Rachtanapun, P. Jantrawut, S. Thanakkasaranee, G. Kasi, J. Tantala, P. Panraksa, and T. Chaiwarit vii

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Contents

Carboxymethyl Chitosan-Based Derivatives in Diagnosis and Targeting . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 205 Kiran Joy, D. Sathya Seeli, and M. Prabaharan Carboxymethyl Chitosan for Drug and Vaccine Delivery: An Overview . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 225 Selin Parmaksız and Sevda S¸enel Carboxymethyl Chitosan and Its Derivatives in Tissue Engineering . . . 257 Fernanda G. L. Medeiros Borsagli Functionalized Carboxymethyl Chitosan Derivatives in Wound Healing . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 281 Erkan T. Baran

Adv Polym Sci (2024) 292: 1–18 https://doi.org/10.1007/12_2023_148 © The Author(s), under exclusive license to Springer Nature Switzerland AG 2023 Published online: 25 April 2023

Introductory Aspects of Carboxymethyl Chitosan Derivatives Irine Rose Antony and R. Jayakumar

Contents 1 Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2 2 Preparation and Physicochemical Characterization of CMC Derivatives . . . . . . . . . . . . . . . . . . . 3 2.1 Preparation of CMC Derivatives . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3 2.2 Characterization of Various CMC Derivatives . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5 3 Properties of CMC . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6 3.1 Physicochemical Properties of CMC . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6 3.2 Biological Properties of CMC . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7 4 Applications of CMC . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 9 5 Conclusion . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 13 References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 14

Abstract Chitosan is a biopolymer produced by chitin deacetylation. Since the amino groups in chitosan are primarily protonated when pH 6.5. In order to overcome the poor solubility in aqueous conditions, chemical modifications are required. Chemical modification of chitosan into carboxymethylated chitosan (CMC) derivatives significantly promotes chitosan’s solubility in a broad pH range. Diverse CMC derivatives can be synthesized by controlling the reaction conditions. In comparison to chitosan, CMC derivatives exhibit better water solubility, moisture retention ability, biocompatibility, biodegradability, antibacterial property, antioxidant activity, and increased metal ion sorption capacity. This chapter delivers an overview of the status and current challenges of different CMC

I. R. Antony and R. Jayakumar (✉) Polymeric Biomaterials Lab, School of Nanosciences and Molecular Medicine, Amrita Vishwa Vidyapeetham, Kochi, India e-mail: [email protected]

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derivatives in antimicrobial, wound healing, hemostasis, anticancer, drug delivery, and tissue engineering areas. Keywords Anticancer · Antimicrobial · Antioxidant · Carboxymethylation · Chitosan · Drug delivery · Nanotherapeutics · Tissue engineering

1 Introduction Chitosan is a natural polycationic biopolymer discovered by Professor C. Rouget in 1859 by N-deacetylation of chitin from crustaceans. Chitosan deacetylation degree varies and usually it is over 50%. Chitin is a linear polymer made up of N-acetyl-β-D-glucosamine and β-D-glucosamine monomer units bound together by β-(1, 4) glycoside bonds [1]. Chitin has rigid crystalline structure with interand intra-molecular hydrogen bonds which makes it insoluble in common solvents [2]. During deacetylation process a portion of the acetyl groups is converted into amine to enhance the influence of primary amine groups and lower the crystallinity of chitosan when compared to chitin. The greater affinity of primary amine groups affects inter- and intra-molecular hydrogen bonds and generate amorphous regions with higher swelling capacities within the biopolymer [2]. Many positively charged amino groups are present in the surface of chitosan and can interact with biomolecules like DNA, phospholipids, proteins, etc. It is the major nitrogen source accessible to living organisms and also has good biocompatibility, biodegradability, and wound healing properties [3]. Even though the chemical structure of chitin and chitosan is differing very little, their chemical reactivity and physical properties are very different. Solubility, chemical reactivity, biodegradability, physiological activities, etc., are influenced by degree of substitution, acetylation degree, and molecular weight. Practical utilization of these materials is limited by their solubility in common solvents. Insolubility of chitosan in alkaline pH limits its application to acidic pH. Chitosan substituted with alkyl chains in adequate length exhibits hydrophobic characteristics. These hydrophobic interactions are influenced by the polymer concentration, temperature, ionic strength of the medium, and the nature and number of hydrophobic sites [4]. Partial N-acetylation or chemical modifications like sulfonation, PEG grafting, quaternization, carboxymethylation, etc., can be employed to maximize its solubility [5]. Recently many CMC derivatives have been developed for biomedical applications. Introduction of carboxymethyl groups in the chitosan polymer can promote its solubility in acidic, basic, and neutral aqueous solutions with less toxicity [6]. Carboxymethyl derivative of chitosan contains both -COOH and –NH2 groups along with polymer chain which renders it insoluble near its isoelectric point [5]. A number of variables, including the reaction media, chemical pathways, degree of deacetylation (DD), degree of substitution (DS), molecular weight, structure, etc.,

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affect how different types of CMCs are synthesized [7]. pKa value of carboxyl group is 4.5 which helps them to be protonated at pH ≥ 7 making carboxymethyl chitosan water soluble in neutral and alkaline pH [8]. In addition to solubility CMC also shows various other desirable physicochemical and biological properties. Molecular weight (MW), number of amine groups, solution pH, solution concentration, and deacetylation degree (DD) primarily influence the antibacterial effectiveness of CMC [9]. Latest results revealed that increased amount of quaternized substitution in CMC lowers the molecular weight and enhances the antibacterial activity of quaternized CMC [10]. It has been discovered that CMC-based bioactive materials can be cross-linked using metal ions, metal oxides, or metallic nanoparticles to enhance their antibacterial activity [11]. CMC serves as a delivery vehicle for bioactive agents like DNA/genes, proteins, anti-inflammatory, antimicrobial, and anticancer medicines [12]. The primary carrier platforms using CMC derivatives based polyelectrolyte complexes, hydrogels, scaffolds are used to release drug adjuvants [13]. Multifunctional wound dressings based on CMC with good solubility, antibacterial/antioxidant characteristics, and biocompatibility have been developed to promote wound healing [14]. Recently, numerous studies shown the advantage of CMC’s simplicity of processing of scaffolds, nanofibers, membranes, hydrogels, composites, and nanoparticles [15]. CMC-based materials are gaining popularity as scaffolds for bone tissue development to encourage osteoblast adhesion and proliferation. In addition, CMC also helps in chondrocyte development and differentiation for cartilage tissue regeneration [16]. In brief, this review outlines the biological and physical characteristics of CMC derivatives as well as their most recent uses in a number of biomedical fields.

2 Preparation and Physicochemical Characterization of CMC Derivatives 2.1

Preparation of CMC Derivatives

Carboxymethyl chitosan (CMC) was first reported by Muzzarelli in 1982. It was prepared by reductive alkylation and direct alkylation methods. In reductive alkylation, N-carboxymethyl chitosan is produced when the -NH2 group of chitosan reacts with the carbonyl group of glyoxylic acid. This reaction is followed by hydrogenation using either NaBH4 or NaCNBH3. This method clearly only places the carboxymethyl substituent on the N-atom and leaves out the O-substitution. However, even under mild conditions, di-substituted N-carboxymethyl chitosan (N, N-diCMC) is formed partially along with N-carboxymethyl chitosan due to the high reactivity of aldehyde. As a result, a significant portion of "N-carboxymethyl chitosan" is di-substituted. Ratio of amine group present in the chitosan is determining the mono/di-carboxymethyl substitution in the glucosamine unit [17].In the

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Fig. 1 Structure of various CMC derivatives like N,O-Carboxymethyl Chitosan (N,O-CMC), NCarboxymethyl Chitosan (N-CMC), O-Carboxymethyl Chitosan (O-CMC), N,N-DiCarboxymethyl Chitosan (N,N-DCMC), O-Succinyl Chitosan (O-SCS), N-Succinyl Chitosan (N-SCS), N-Carboxybutyl Chitosan (N-CBCS), and N-Carboxyethyl Chitosan (N-CECS)

direct alkylation approach, N- and O-carboxyalkyl chitosan derivatives are made using monohalocarboxylic acids and monochloroacetic acid under various reaction conditions. The degree of substitution and N vs O selectivity in carboxyalkylation are determined by the reaction conditions. Chitosan must first be stimulated by immersing it in an alkaline solution before carboxymethylation can occur in solvent of water/isopropyl alcohol. Solely the amine groups are stimulated during the carboxymethylation of chitosan in the mild alkaline medium (pH 8–8.5) leading to N-substitution in the presence of monochloroacetic acid [18]. All of the chitosan molecules in solution can be mono- or di-N-substituted despite the fact that the chitosan that precipitated at this pH was redissolved gradually as the reaction moves forward. Nevertheless, mixed N- and O-CMC derivatives can be produced by alkylation using monochloroacetic acid with replacement at the C6 and C3 -OH groups and additional substitution into C2-NH2 position [1]. Different CMC derivatives (Fig. 1) can be produced by selecting the proper reaction conditions and reagents. Researchers have created derivatives which are soluble in alkaline and acidic solutions through the carboxymethylation of the

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chitosan structure. Amino and hydroxyl groups found in its chains operate as the reactive sites for carboxymethylation of chitosan. While the reaction is conducted at room temperature or in a cold bath, in an isopropanol/water solution with monochloroacetic acid and NaOH, (O-CMC) is mostly produced. However, higher reaction temperatures yield (N-CMC) and (N,O-CMC). Instead, by reacting chitosan with glyoxylic acid, followed by reduction with NaCNBH3, it is possible to create NCMC and N, O-CMC [17].

2.2

Characterization of Various CMC Derivatives

It has been difficult to reliably characterize biopolymers and their derivatives. Unstable biopolymer solutions provide unreplicable outcomes and incorrect descriptions of macromolecular characteristics. To more accurately characterize biopolymer solutions like CMC, experimental techniques should be devised for addressing stability issues, effects of stable aggregates, and other problems [19]. Fourier Transform Infrared (FTIR) & Nuclear Magnetic Resonance (NMR) spectroscopy, differential scanning calorimetry, X-Ray Diffraction (XRD), and capillary zone electrophoresis are the most common techniques used to characterize CMC. O- and N-carboxymethylation can be obtained under carefully regulated reaction conditions. Nuclear Magnetic Resonance was used to calculate the substituent yield at different locations [20]. There is not much information on Sample Light Scattering (SLS) or multi-detector Gel Permeation Chromatography (GPC) characterization of CMC. Pullulan or dextrans were used as a calibration standard to measure the molecular weight distributions (MWD) of sulfated and carboxymethylated chitosan [19]. MWD of O-CMC with varying levels of acetylation were established [21]. O-carboxymethylated chitosan was also investigated using Size Exclusion Chromatography coupled with Multi-Angle Light Scattering (SEC-MALS) in 0.1 M NaCl and it was found that the samples were heterogeneous and had bimodal distribution [19]. FTIR analysis for O-CMC showed the representative peaks of chitosan at 1,560 cm-1 (-NH bending of amine-I and amide II), 1,650 cm-1 (-C=O stretching amide-I), 1,030 and 1,080 cm-1 (C - O stretching), 1,150 and 896 cm-1 (-C - O C), 1,320 and 1,260 cm-1 (-CN) and also the characteristic peaks of O-CMC at 1,630 cm-1(antisymmetric -NH3+ stretching) and 1,730 cm-1(-COOH) [22]. The proton assignment of CS can be seen in the proton NMR spectrum together with the distinctive proton signal of O-CMC, which appears in the range of 4.0–4.1 ppm, indicating that the carboxymethyl groups are attached to the CS [5]. IR spectrum of N-CMC showed characteristic peaks at 1,527 (-NH2), 1,409 (-CH2 in-CH2OH), 1,720 (-C=O), 727 (pyrene ring skeleton), and 1,303 cm-1(-CH3 in -NHCOCH3) [20]. The distinctive peak of N,O-CMC emerged at an angle of 32.12226° on the XRD, which showed less intense than the original peak of CS.

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3 Properties of CMC 3.1

Physicochemical Properties of CMC

The degree of carboxymethylation and modifying the reaction conditions had a significant improvement on the solubility of CMC in water [23]. Recent research by Chen and Park on the solubility of O-CMC with varying degree of substitution (DS) revealed that variable reaction temperatures and solvents led to diverse O-CMC water solubilities [5]. The optimal DS value of CMC is 0.4–0.45. At this range CMC dissolves in water [23]. The investigation also showed NaOH concentration had an impact on the DS of the resulting CMC and recommended that the best alkali concentration for the carboxymethylation process appeared to be 50% NaOH solution. The hard crystalline structure of chitosan was not sufficiently broken by the low NaOH concentration, which decreased the monochloroacetic acid’s ability to permeate the interlocking polymer chains. A different study [24] showed that the ratio of NaOH to monochloroacetic acid in the reaction mixture had a significant effect on the aqueous solubility of CMC. Higher concentrations of alkali (above 60%) upheld side reactions between monochloroacetic acid and NaOH thereby reducing the concentration of monochloroacetic acid [25]. Over the entire pH range of 2–12, the CMCs were either insoluble or slightly soluble at low NaOH to monochloroacetic acid ratios (0.28–0.40). However, with larger NaOH to monochloroacetic acid ratios, products with optimal reaction time were produced and showed better water solubility [24]. The different mass ratios of chloroacetic acid to chitosan at 5:1, 8:1, and 10:1 in the presence of microwave irradiation conditions were investigated, and the findings showed that after 20 min, the DS increased with the increased ratio from 5:1 to 8:1, however that at ratio of 10:1, showed no further increase [25]. At different pH levels, water insolubility of CMC varied according to the DS. The insoluble area may be the result of either amide production upon thermal drying or agglomeration of highly acetylated chain segments. An electrostatic repulsion between -COO groups on the particles, intermolecular H-bonding of CMC, and hydrophobic contact between the hydrophobic groups in the CMC (such as acetyl groups and glucosidic rings) are the driving forces behind the aggregation behavior that is observed in neutral, diluted solutions of O – CMC. According to the reported study, O-CMC’s critical aggregation concentration lies between 0.042 and 0.05 mg/mL [26]. Furthermore, it has been noted that monocarboxymethyl chitosan with 87–90% DS possesses polyampholytic properties, allowing the production of transparent gels or solutions based on the polymer content at alkaline and neutral pH values, but clusters under acidic conditions. Moisture retaining capabilities of CMC have drawn a lot of interest due to its potential uses in clinical medicine, cosmetics, and other biomedical fields (Fig. 2) [27]. It has been reported that 0.25% aqueous solution of CMC and 20% aqueous solution of propylene glycol were similar in terms of moisture retention capacity, and its viscosity was nearly identical to that of hyaluronic acid, a substance with

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Fig. 2 Properties of CMC

well-known moisture retention capabilities [28]. CMCs have demonstrated outstanding adsorption properties. This may be due to strong hydrophilicity, abundance of hydroxyl and amino groups, adaptable polymer chain structure, and ability to take on the proper configuration for complexation with metal ions [29]. Two theories have been proposed for the interpretation of the mechanisms underlying the absorption of various metal ions. It has been found that through inter- or intra-molecular complexation, metal ions are bound to multiple amine groups from the same chain or from distinct chains in the bridge model, whereas in the pendant model, they are bound to a single amine group [30].

3.2

Biological Properties of CMC

CMC derivatives exhibit antimicrobial properties. The bactericidal effects of chitosan and different CMC derivatives on E. coli follow the order N,O-CMC< chitosan7 days) have been shown to be resistant to 500–5,000 times the doses of antibiotics and chemical biocides required to kill the same organism’s free-floating (planktonic) cells [33]. Coating of CMC on magnetic iron oxide nanoparticles could dissolve biofilms and showed bactericidal effect against E. coli and S. aureus. Additionally, CMC-magnetic nanoparticles (MNPs) effectively entered into S. aureus and E. coli biofilms with the help of magnetic field, and after 48 hours of incubation, it was able to significantly reduce the viability of the bacteria compared to respective controls [34]. Intermolecular non-covalent interactions like hydrogen bonds, host–guest complexation, and ionic interactions hold two or more molecular entities together to form soft materials called supramolecular hydrogels [35]. In comparison to chitosan hydrogels, CMC supramolecular hydrogels exhibit better water solubility, moisture retention ability, biocompatibility, biodegradability [36], antibacterial property, antioxidant activity [37], and increased metal ion adsorption capacity [38] due to its higher chelation and chain flexibility.

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CMCs have other advantageous traits like anticancer and anti-tumor properties. CMC exhibits inhibitory effects on hepatic tumor cell BEL-7402 migration in vitro to highly prevent lung metastasis associated with hepatoma-22 in Kunming mice. For oral delivery of the anticancer medication paclitaxel (PTX), an amphiphilic CMC-quercetin (CQ) compound was created in an effort to increase the drug’s oral bioavailability by making it more water soluble and avoiding the P-glycoprotein (P-gp) drug efflux pumps [39]. Although most organisms possess antioxidant protection systems that safeguard them from oxidative stress, damage cannot be completely prevented. According to reports, amino and hydroxyl groups present inside the polymer chains allow CMC to exhibit sufficient antioxidant activity. Additionally, it was discovered that lowering the molecular weight of CMC causes an increase in antioxidant activity, which can be related to the less intra- and intermolecular hydrogen bonds. Low molecular weight CMCs produced by oxidative degradation were examined for their superoxide anion scavenging abilities, and it was discovered that they had a higher activity [40]. Chitosan can be used as a powerful antifungal agent to prevent C. albicans infections. It was discovered that CMC outperformed chitosan in terms of antifungal activity [41].

4 Applications of CMC CMC has distinct physical, chemical, and biological properties with better biocompatibility, and has attracted significant interest in a wide range of biomedical applications (Fig. 4), including bioimaging, drug delivery, hemostasis, tissue engineering, bacteriostatic agents, and blood anticoagulants. It has also been shown that numerous -COOH and –NH2 groups in CMC are responsible for the material’s superior pH tolerance and ion sensitivity in aqueous conditions [42]. Jayakumar et al. detailed about the metal uptake (Ni2+, Zn2+, and Cu2+) performance and bioactivity of CMC-graft-d-glucuronic acid for environmental and tissue engineering applications. By selecting different types of side chains, grafting d-glucuronic acid onto CMC provides desirable qualities and broaden the range of potential applications for chitosan [43] CMCs can be used as carrier matrix in encapsulation system. Due to its hydrophobic nature, chitosan has less encapsulation efficiency than CMC. The reaction conditions for the preparation of CMC can be optimized to encapsulate various hydrophilic drugs [13]. Recently significant advancements have been made on the use of CMC-based polymers for anticancer medication delivery. Prolonged release of anticancer medications, injectable hydrogels made of N,O-CMC with multi-aldehyde guar gum (MAGG) were utilized. The findings demonstrated that the hydrogel exhibited exceptional self-healing, mechanical, biocompatible, and thixotropic properties in addition to excellent ability to destroy breast cancer cells (MCF-7) [44]. Hydrogels are 3D porous hydrophilic polymeric systems that replicate natural tissue by having a high water content [45]. Hydrogels are utilized as wound dressings because of their biocompatibility, extensibility, and elasticity. Hydrogel’s

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Fig. 4 Applications of CMC

main flaw is that they are more likely to deform and crack as a result of outside forces, which makes it difficult for them to act at the site of action. A novel improvement over brittle hydrogels is the use of self-healing adherent hydrogels [46]. These hydrogels have the innate ability to repair themselves after being damaged, regaining their structural and functional capabilities and mimicking the characteristics of real tissue. An in situ AgNps containing N, O-CMC self-healing hydrogel with carboxyl-metal-ligand co-ordination bond exhibiting adhesive, conductive, antibiofilm, and antibacterial properties were reported. The reversible non-covalent interaction of the polymeric network promotes the development of hydrogels with effective self-healing capabilities as well as structural integrity and mechanical strength that are beneficial to be employed as tissue sealant for infected wounds [47]. Blood is the most significant fluid connective tissue in the human body and has a variety of functions in maintaining healthy physiological processes, including gas and nutrition movement, immunological surveillance, and hemostatic responses [48]. Blood loss causes a number of issues, including death. The first objective in treating a significant bleeding wound is to stop the bleeding as soon as possible [49]. Although artificial dressings have effective hemostatic and antibacterial qualities, the possibility of residual raw materials restricts their use [50]. Natural polymers like cellulose [51], alginate [52], chitosan [53], and gelatin [54] receive greater attention in bleeding treatment than synthetic polymers due to their beneficial qualities, such as absorption capacity, biocompatibility, and ease of processing. In comparison to chitosan, CMC offers improved biocompatibility and biodegradability as well as a higher solubility in neutral conditions and a low swelling degree

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[55]. So it is better suited than chitosan for practical hemostatic applications. Sponges with exceptional absorption capabilities are frequently utilized for hemostasis during an emergency. Hemostatic sponge developed on modified Graphene Oxide (GO) and CMC exhibiting outstanding mechanical strength and water absorption capabilities has been reported. Additionally, hemostasis tests revealed that the composite sponge had a good hemostatic efficacy [56]. Creation of new hemostatic drugs with different mechanisms of action from those of more established ones is still a difficult task. Self-assembling peptide hydrogels have become a new hemostatic agent due to their intrinsic biocompatibility, biodegradability, and designability, among other factors. Recently, Hao et.al synthesized a hemostatic agent consisting of self-assembling short peptide and O-CMC. The transglutaminase catalyzed reaction happening between the ultra-short peptide and O-CMC resulted in the production of intra-molecular-(-glutamyl)-lysine isopeptide bonds, which in turn facilitates the development of flexible and intertwined nanofibers. This provides a potential strategy for making alternate hemostatic materials [57]. Nanoparticles can lower drug toxicities and regulate the release of encapsulated or related pharmaceuticals. So chitosan/O-CMC nanoparticles (CS/CMC-NPs) can be used to promote the bioavailability of the anticancer drug during oral delivery [58]. Novel bacteriostatic Ag/AgO/CMC and Ag/AgO/CMC/Aspirin (ASP) hydrogels were developed under mild conditions employing AgNO3 as a crosslinking agent. These hydrogels are good and efficient drug delivery carriers with many applications in the treatment of pain. It has the benefits of enhancing the cumulative release of ASP in the intestinal location and boosting bacteriostatic activity [33]. Due to their excellent biocompatibility, biodegradability, hemostatic and antibacterial qualities, natural biopolymers play a crucial role in wound healing. Additionally, they offer a moist environment at the wound site to aid in the healing process [33]. Because CMC chains contain amino groups, they can interact with other polymer’s aldehyde functional groups to form Schiff base bond that is necessary for self-healing [59]. Carboxymethylated cotton exhibits superior hemostatic capacity than cotton fabric and can be used as wound dressing. The cotton fabric that had been modified (MCF-0.39) by carboxymethylation had good water absorption and swelling properties [60]. To maintain their dimensional integrity for potential usage in wound care applications, CMC was cross-linked using a microwave method [35]. A CMC-based hydrogel enhanced with modified cellulose nanocrystals to promote deep partial thickness burn wound healing has also been reported [61]. For use as wound dressings, CMC-polydopamine hydrogels from fungal mushrooms (FCMCS-PDA) with tissue adhesive/hemostatic/self-healing/antibacterial capabilities have been produced. On E. coli and S. aureus, the PDA-containing hydrogels exhibit good antibacterial capabilities. Moreover, the combination of FCMCS and PDA produced rapid blood coagulation within 60 s. Therefore, this multifunctional adhesive hydrogel has great promise in infected wounds treatment [62]. Carboxymethyl chitosan-grafted poly(vinylpyrrolidone-iodine) (CMC-gPVPI) microspheres to promote chronic wound healing were also reported recently

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[63]. To aid in the healing of burn wounds, a CMC-based hydrogel that is loaded with curcumin (Cur) and cross-linked by a ROS-sensitive linker was created. Attributed to the presence of ROS-sensitive cross-linkers, this hydrogel can remove excess ROS from the microenvironment of the burn wound site [64]. Currently, nanotherapeutics and nanoparticle mediated drug delivery for cancer therapy are popular topics because they can get beyond the drawbacks of traditional drug delivery systems [65]. The delivery of anticancer drugs via nanoparticles that are functionalized with monoclonal antibodies has tremendous potential. A number of cancer cells have been shown to overexpress the human EGF receptor tyrosine kinase known as the epidermal growth factor receptor (EGFR) [66]. In order to deliver paclitaxel (PTXL) to cancer cells that overexpress the epidermal growth factor receptor (EGFR), a targeted drug delivery system made of cetuximab (Cet) conjugated O-carboxymethyl chitosan (O-CMC) nanoparticles has been developed. In comparison to non-targeted nanoparticles, these targeted nanoparticles had greater anticancer activity, and the nanoformulation caused accelerated cell death (verified by flow cytometry) because of their higher cellular absorption [67]. In order to create regulated medication delivery systems, N-CMC beads have been created. They were made utilizing the tripolyphosphate counter polyanion ionotropic gelation technique. It was discovered that N-CMC beads were pH sensitive. Drugs were released from N-CMC beads more quickly in simulated intestinal fluid (pH 7.4) than in simulated gastric fluid (pH 1.4). These considerations imply that when compared to CS beads, N-CMC beads may be an effective controlled drug delivery device for oral administration because they prevent the release of drug at extremely acidic gastric fluid in the stomach [68]. Recently, it was discovered that the widely used diabetes medication metformin inhibits the growth of pancreatic cancer by inhibiting mTOR (mammalian target of rapamycin) and activating AMP kinase (AMPK) [69]. Metformin’s use as a fullfledged anticancer medicine is now hampered by its limited absorption and brief half-life [70]. O-CMC has attracted a lot of interest in drug delivery applications. Its reactive O-carboxymethyl substitution may interact with NH3+ group of metformin, making it easier to incorporate during the production of nanoparticles. In vitro, the produced NPs with spherical morphology and a size of 240 nm showed pH-sensitive metformin release. According to cytotoxicity experiments, the drug-incorporated NPs significantly harmed pancreatic cancer cells (MiaPaCa-2) in comparison to healthy L929 cells [71]. Folate linked carboxymethyl chitosan-zinc sulfide nanoparticles were developed for cancer cell imaging and drug delivery. The targeting of nanoparticles into cancer cells with the help of CMC and folic acid conjugation increases the bioavailability of 5-FU. Because it employs controlled drug administration via CMC and uses fluorescence from a ZnS:Mn quantum dot to visualize the progression of this drug carrier system, this study is noteworthy in the area of anticancer drug delivery [72]. Turmeric, the rhizome of the plant Curcuma longa, is the source of the hydrophobic polyphenol known as curcumin. Numerous research on curcumin have identified low absorption, fast metabolism, and elimination of curcumin as the primary causes of this polyphenolic compound’s poor bioavailability to humans

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[73]. The hydrophobic anticancer medication curcumin was put into a nanoformulation that was built on O-CMC. With this method, curcumin was first solubilized and then stabilized in O-CMC nanoparticles. Drug-loaded biocompatible nanoparticles with improved cellular absorption were easily obtained. They exhibited anticancer effects comparable to those of free curcumin [74]. Antibacterial activity of carboxymethyl derivatives such as O-carboxymethyl chitosan (O-CMC), N,O-carboxymethyl chitosan (N,O-CMC), and Ncarboxymethyl chitosan (N-CMC) nanoparticles was studied. Through straightforward chemical processes, these carboxymethylated nanoparticle derivatives have been developed for use in a variety of medical applications. According to antibacterial studies, CS nanoparticles were less effective against bacteria than OCMC and N, O-CMC nanoparticles. Out of the three, N,O-CMC nanoparticles demonstrated the strongest antibacterial activity, and no colonies were discovered in comparison to the control [75]. Numerous ailments, ranging from mild skin infections to life-threatening disorders, are caused by S. aureus. It is an extracellular pathogen and can occasionally induce intracellular infections where it can penetrate both professional and non-professional phagocytes [76]. Because antibiotics only partially penetrate macrophages and epithelial cells, intracellular bacterial infections are chronic, recurring, and challenging to cure. Maya et.al created biocompatible tetracycline-entrapped OCMC nanoparticles (Tet-OCMC Nps) for the sustained administration of Tet into cells. Tet-O-CMC Nps bind to and aggregate with S. aureus, causing an increase in medication concentrations at the site of infection. Tet-O-CMC Nps proved to be a useful nanoformulation to treat intracellular S. aureus infections by being six times more effective than Tet alone in killing intracellular S. aureus in differentiated THP1 macrophage cells and HEK-293 cells [77].

5 Conclusion This chapter provides a brief overview about the perspectives of CMC in terms of synthesis, properties, and various biomedical applications of different carboxymethyl chitosan derivatives. The stiff crystalline form of chitosan causes it to have poor solubility in water, which prevents it from being used effectively in a variety of operations. The solution to this issue is to prepare its chemically modified water-soluble derivatives like CMC which imparts improvement of physicochemical and biological properties making them a promising candidate in various biomedical applications. Due to the enhanced physicochemical properties of CMC, a wide range of their hydrogels, composites, complexes, and conjugates have been created for use in drug/gene therapy, tissue engineering, controlled therapeutic release, and other applications. The latest outcome of CMC derivatives used in a variety of biomedical fields, including drug delivery, hemostasis, tissue engineering, regenerative medicine, and wound healing were covered in this review. The excellent antibacterial and hemostatic action makes CMC particularly suitable for wound healing. It is noted

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that the majority of CMC applications are still in the laboratory stage and need additional research in order to be used in a clinical setting as commercial biomedical and pharmaceutical materials.

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Adv Polym Sci (2024) 292: 19–30 https://doi.org/10.1007/12_2023_152 © The Author(s), under exclusive license to Springer Nature Switzerland AG 2023 Published online: 27 July 2023

Preparation of Different Types of Carboxymethyl Chitosan Derivatives Irine Rose Antony and R. Jayakumar

Contents 1 Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2 Preparation of Different Types of Carboxymethyl Chitosan Derivatives . . . . . . . . . . . . . . . . . . . 2.1 Synthesis of O-Carboxymethyl Chitosan (O-CMC) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2.2 Synthesis of N-Carboxymethyl Chitosan (N-CMC) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2.3 Synthesis of N,O-Carboxymethyl Chitosan (N,O-CMC) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2.4 Synthesis of N, N-Dimethyl Carboxymethyl Chitosan (N, N-dCMC) . . . . . . . . . . . . . . . . 2.5 Synthesis of N-Carboxybutyl Chitosan (N-CBC) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2.6 Synthesis of N-Carboxyethyl Chitosan . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2.7 Synthesis of N-Succinyl Chitosan . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2.8 Synthesis of O-Succinyl Chitosan . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3 Conclusion . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

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Abstract Chitosan (CS) has long been regarded as a substance with numerous exceptional qualities, including biodegradability, biocompatibility, non-toxicity, and ecological sustainability. However, its widespread use is hampered by its weak solubility in water and in typical organic solvents. This limits its applications in biomedical and other areas. Chemical modification of chitosan improves its solubility in aqueous solutions. The use of carboxymethyl moieties to chemically modify the hydroxyl and amino groups already present on the CS chains would increase the solubility of chitosan and also endow it with a variety of special chemical, physical, and biological properties. The water solubility, antibacterial, absorption/retention, and emulsion stabilizing properties of the carboxymethyl chitosan (CMC) compounds have all been improved. They can engage with cells to promote cell proliferation, regeneration, and wound healing. So, this review will I. R. Antony and R. Jayakumar (✉) Polymeric Biomaterials Lab, School of Nanosciences and Molecular Medicine, Amrita Vishwa Vidyapeetham, Kochi, India e-mail: [email protected]

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focus on different derivatives of CMC with special emphasis on their preparative methods. Keywords Biomedical applications · Carboxymethylation · Chemical modification · Solubility · Succinyl chitosan

1 Introduction Chitosan is the most prevalent carbohydrate found in nature. However, applications of chitosan are severely constrained by its extremely solid crystalline structure, which is created by powerful hydrogen bonding that renders it insoluble at neutral or alkaline pH. Only aqueous solutions with a pH of less than 6.5 exhibit solubility [1]. Numerous water-soluble derivatives have been made by quaternarization [2] adding hydrophilic groups like dihydroxyethyl, hydroxypropyl, sulfate [3, 4], hydroxylalkylamino [5], phosphate [6], carboxyalkyl [7] groups like carboxyethyl, carboxymethyl or carboxybutyl [8] to the macromolecular chain of chitosan. Chitosan that has undergone carboxymethylation has enhanced water solubility, which aids in overcoming this obstacle [9, 10]. In comparison with other water-soluble chitosan derivatives, CMC has received the maximum attention because of its simplicity in production, ampholytic nature, and wide range of potential applications. At a pH of 7, CMC is soluble in water. Along with other beneficial qualities including biocompatibility, biodegradability, biological activity, and low toxicity, it has a high viscosity, a big hydrodynamic volume, and the capability to generate films and gel [11]. It has been reported that the threshold degree of substitution (DS) value that triggers CMC to readily dissolve in water is between 0.4 and 0.45 [8]. The amount of NaOH in the reaction influenced DS of the resultant CMC and 50% NaOH solution was the optimal concentration of alkali for the carboxymethylation process [12]. It was also found that electrostatic repulsion, intermolecular H-bonding, and hydrophobic interaction in the CMC molecules causing aggregation in neutral and diluted aqueous formulations [13]. Direct alkylation and succinylation of chitosan can produce different types of carboxymethyl chitosan derivatives [14, 15]. Water-soluble CMC derivatives are very useful in different types of biomedical [15, 16] and environmental applications. So, this review will focus on preparatory methods of various CMC derivatives and mechanisms involving in their preparation.

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2 Preparation of Different Types of Carboxymethyl Chitosan Derivatives 2.1

Synthesis of O-Carboxymethyl Chitosan (O-CMC)

O-CMC is an amphiprotic ether analog that has -NH2 and -COOH groups within the molecule. A highly alkaline media is used for the preparation of O-CMC. It can be made by adding chitosan to NaOH in a reaction container with isopropanol and then stirring for 1 h at room temperature. The reaction mixture is then added dropwise for 30 min while being diluted in isopropanol with monochloroacetic acid. The whole mixture is held at 55°C for the 4 h of reaction. Then, the product is filtered, rinsed with ethyl alcohol and then vacuum dried [17]. According to experimental evidences, the reaction conditions and level of carboxymethylation affect the O-CMC solubility in water. When O-CMC is prepared at 0–10°C, it is water soluble. However, at a neutral pH, the O-CMC produced at 20–60°C remains insoluble in water. As the water to isopropanol ratio in the reaction mixture increased, the amount of carboxymethylation as well as insolubility at higher pH decreased. On the contrary, a spike in reaction temperature amplified the carboxymethylation degree and insolubility at lower pH [18]. The preparation method of O-CMC is shown in Fig. 1. A more effective alternative to currently existing eye drops for the treatment of glaucoma consisting of O-CMC nanoparticles incorporated with dorzolamide has been studied. It reduces pulse entry while providing personalized medication release and increased bioavailability [19]. O-CMC-based magnetic nanogel was developed for targeted and sustained delivery of antitumor drug. The presence of hydrophilic amino and carboxyl groups in the O-CMC helps nanogel to exhibit fast swelling and prolonged drug release [20].

2.2

Synthesis of N-Carboxymethyl Chitosan (N-CMC)

N-CMC was prepared by reduction of the in situ produced imine generated by reaction of amine group of chitosan with the aldehyde group of glyoxylic acid. The reducing agent can be either NaBH4 or NaBH3CN. However, this technique

Fig. 1 Preparation of O-CMC

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Fig. 2 Synthesis of N-CMC

involved two steps [14]. The preparation method is shown in Fig. 2. Alternatively, N-CMC was also developed by dissolving chloroacetic acid in water. After that, pH of the mixture was brought to 7.0 by adding NaOH. After that chitosan was mixed into the aforementioned solution and vigorously stirred at 90°C for 4 h to obtain NCMC [21]. Below 80°C, this reaction developed very slowly, and little product was produced. However, the product became yellow at 95°C. Consequently, the ideal temperature was 90°C. The pH of the reacting system should be kept at 7.0 by adding 20% Na2CO3 to consume the HCl produced during the N-CMC process. Then filtrate was diluted using 95% ethanol (v/v 1:3) to remove the unreacted chitosan [21]. Felicio et al. provided a description of spherical aggregation behavior of N-CMC. Images from AFM reported the static and dynamic light scattering behavior [22]. Given its prolonged moisturizing impact on the skin, N-CMC is a beneficial functional constituent of cosmetic agents. Film-forming potential of NCMC helps in maintaining the smoothness of skin [23].

2.3

Synthesis of N,O-Carboxymethyl Chitosan (N,O-CMC)

The amine and 6-hydroxyl positions of certain glucosamine units are modified by carboxymethyl substituents in the chitosan to obtain N, O-CMC. It is water soluble and has specific physical and biological characteristics which includes gel formation, moisture retention, and good biocompatibility. N,O-CMC is an excellent biomaterial due to these properties [24]. N, O-CMC can be made by treating chitosan with isopropyl alcohol and NaOH to make an alkaline slurry. Into this slurry monochloroacetic acid was added with a subsequent increase in temperature(50°C) and vigorous stirring (Fig. 3). Then the reaction mixture was filtered and vacuum dried to obtain pure N,O-CMC [24, 25]. Topical treatment of dialdehyde crosslinked N,O-CMC hydrogel significantly reduced the postoperative fibrosis without adverse cardiac side effects in a rabbit surgical sternotomy pericardial adhesion model [26]. Curcumin and 5-fluorouracil loaded N,O-CMC nanoparticles have been developed for the management of colon cancer. The formulation exhibited extended drug release profile, improved plasma concentrations, and enhanced anti-cancer effects [27]. By graft copolymerization of 1-vinyl-2-pyrrolidone and sodium acrylate with N,O-CMC, a novel superabsorbent

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Fig. 3 Preparation of N,O-CMC

Fig. 4 Preparation of N,N-dCMC

polymer was developed. Poly(1-vinyl-2-pyrrolidone) (PVPD) segments were also added to the N, O-CMC to increase their water affinity to alter their haloduric properties [28]. A self-healing hydrogel with in situ silver nanoparticles (AgNps) and N,O-CMC was developed. This self-healing hydrogel showed better antibacterial activity against S. aureus and E. coli [29]. N,O-CMC is also a good choice for transporting hydrophobic drugs like curcumin [30]. Due to their aromatic structure, dye effluents are biologically non-degradable and challenging to handle. Adsorption is typically regarded as a useful technique for reducing the amount of solubilized dyes in an effluent. As a potential adsorbent for elimination of Congo Red dye in wastewater treatment, a unique N,O-CMC/montmorillonite (MMT) nanocomposite has been developed and reported. This nanocomposite with a molar ratio of 5:1 of N,O-CMC to MMT had the highest adsorption capacity [31].

2.4

Synthesis of N, N-Dimethyl Carboxymethyl Chitosan (N, N-dCMC)

N,N-dCMC can be made by treating chitosan with glacial acetic acid and glyoxalic acid. Addition of NaBH4 into this reaction mixture resulted in complete dissolution of chitosan to produce N,N-dCMC (Fig. 4) [32]. N, N-dCMC has filmogenic properties and produces transparent films durable under mechanical stress. Additionally, N, N-dCMC is very effective in chelating metal ions. The chelating properties of N, N-dCMC can disrupt physico-chemical behavior of calcium and magnesium salts. Homogeneous interaction of CMC with

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calcium phosphate(CaP) also promotes mineralization of bone tissue. dCMC-CaP exhibits osteoinductive characteristics of chitosan as well as mineralization abilities of amorphous calcium phosphate [32]. N, N-dCMC combined with bone morphogenetic protein (BMP) was used to stimulate the healing of articular cartilage lesions. Results show that BMP-7 promotes in vivo cell proliferation with a chondrocyte phenotype in the articular environment, which results in a partial repair of the articular surface lesions [33].

2.5

Synthesis of N-Carboxybutyl Chitosan (N-CBC)

N-carboxybutyl chitosan (N-CBC) is a water-soluble amphoteric polymer at neutral, basic, and acidic conditions. N-CBC was obtained by combining chitosan and levulinic acid with a reducing agent (sodium borohydride) (Fig. 5) [34, 35]. According to the chemical conditions, the reaction usually produces NCBC or a cyclic compound called 5-methyl pyrrolidinone chitosan (5-MPCs) [36]. Therefore, it is crucial to adequately regulate the reaction media to obtain pure N-CBC. Film-forming ability, solubility, and antimicrobial capacity of N-CBC make it advantageous to be employed in a wide range of applications including wound management, skin care, and cutaneous tissue regeneration [37, 38]. It was shown that N-CBC causes the development of structured repair tissue [39]. In order to encourage organized tissue regeneration, soft pads of N-CBC were applied to donor sites in patients recovering from plastic surgery. Improved histoarchitectural order with vascularization was observed without inflammatory cells at the dermal level compared to control donor sites. In addition, less characteristics of the Malpighian layer production were found at the epidermal level. It indicates that N-CBC promotes the development of cutaneous tissue and lessens anomalous healing [40]. When NCBC was tested against cultures of several pathogens, it exhibited bactericidal, inhibitory, and candidacidal properties. These results suggested that N-CBC is suitable in wound healing [37].

Fig. 5 Preparation of N-CBC

Preparation of Different Types of Carboxymethyl Chitosan Derivatives

2.6

25

Synthesis of N-Carboxyethyl Chitosan

The N-carboxyethyl chitosan (N-CEC) derivative can be made utilizing 3-halopropionic acids in mild alkali conditions (pH 8–9, NaHCO3) (Fig. 6). The first formulation of CEC from chitin and chitosan using 2-chloropropionic acid was reported [41]. In line with this, N-(2-carboxyethyl) chitosan was prepared using low-MW chitosan with 3-halopropionic acids at 60°C in a mild alkaline medium (pH 8–9, NaHCO3) [42]. Biocompatible N-CEC has antioxidative and antimutagenic activity [43]. N-CEC has proven to be an effective medium for introducing nitrogen oxide for therapeutic applications or for the delivery of hydrophilic drug via the skin [44]. Having both amine and carboxyl in its structure, N-CEC can be transformed into ammonium cation and carboxylate anion. Therefore, the negative charges from carboxylate anions and the positive charges from the ammonium cations of N-CEC can bind with bacterial membrane to impart antibacterial properties [45].

2.7

Synthesis of N-Succinyl Chitosan

N-succinyl chitosan (N-SC) is developed by addition of succinyl groups into the glucosamine units of chitosan. Degree of succinylation can be altered by adjusting the quantity of succinic anhydride present in the reaction media. N-SC can be prepared by adding succinic acid (dissolved in acetone) into chitosan which was dissolved in acetic acid (Fig. 7) [46, 47].

Fig. 6 Preparation of N-CEC

Fig. 7 Preparation of N-SC

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In water, N-SC can self-assemble and form nanospheres. Without the need of organic solvents, extreme temperatures, surfactants, or other aggravating factors, the N-SC nanospheres formed spontaneously in relatively moderate conditions [46]. As a drug carrier, N-SC has advantageous qualities like low toxicity, biocompatibility, and longer retention in the body. N-SC structure includes -NH2 and -COOH groups, which make it easily reactive with a variety of substances. The water-soluble carbodiimide and mitomycin (MMC) or an active ester of glutaric MMC might be used to make the water-soluble and water-insoluble conjugates. The MMC and N-SC conjugates demonstrate effective antitumor activity against a variety of malignancies [47]. N-SC alginate hydrogel beads were developed for the regulated distribution of nifedipine drug. At pH 1.5, a small amount of nifedipine was liberated from hydrogel bead (11.6%); however at pH 7.4, release was close to 76%. These findings strongly indicated that the N-SC/alginate hydrogel bead would be a promising pH-sensitive carrier for delivery of drugs inside digestive tract [48].

2.8

Synthesis of O-Succinyl Chitosan

An effective method was reported for preparing water-soluble O-SC. Amine group of chitosan was initially shielded by phthaloyl group, and O-succinylation was then finished (Fig. 8). Hydrazine hydrate was used to finally eliminate the phthaloyl group. The water solubility of chitosan was significantly increased after adding succinyl group to the hydroxyl group. The free amine group of O-SC can serve as a useful intermediate, allowing other chemical modifications for adjusting solubility and biodegradability. Phthaloyl group used as a protecting group for the amino group of chitosan because it has a greater nucleophilicity than hydroxyl group [49]. Like the other chitosan derivatives mentioned above, this water-soluble derivative is also used for various biomedical applications such as wound healing, drug delivery, and skin care [50].

Fig. 8 Preparation of O-SC

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3 Conclusion Preparative methods of different types of CMC derivatives were overviewed. Carboxymethylation was done by direct alkylation, reductive alkylation, and Michael addition methods. The mechanisms and reactions conditions involving in the preparation of CMC derivatives were discussed in detail. All the CMC derivatives are water soluble in comparison with chitosan. Water solubility of this CMC derivatives opens new biomedical, environmental, and other biological applications.

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15. Jayakumar R, Prabaharan M, Nair SV, Tokura S, Tamura H, Selvamurugan N (2010) Novel carboxymethyl derivatives of chitin and chitosan materials and their biomedical applications. Prog Mater Sci 55:675–709 16. Mathew ME, Mohan JC, Manzoor K, Nair SV, Tamura H, Jayakumar R (2010) Folate conjugated carboxymethyl chitosan-manganese doped zinc sulphide nanoparticles for targeted drug delivery and imaging of cancer cells. Carbohydr Polym 80:442–448 17. Biswas S, Ahmed T, Islam M, Islam M S, Rahman M (2020) Biomedical applications carboxymethyl chitosans. Chitin- and chitosan-based polymer materials for various applications, vol 3, pp 433–470 18. Chen XG, Park HJ (2003) Chemical characteristics of O-carboxymethyl chitosans related to the preparation conditions. Carbohydr Polym 53:355–359 19. Shinde U, Ahmed MH, Singh K (2013) Development of dorzolamide loaded 6-Ocarboxymethyl chitosan nanoparticles for open angle glaucoma. J Drug Deliv 2013:562727 20. Demarchi CA, Debrassi A, de Campos BF, Corrêa R, Filho VC, Rodrigues CA, Nedelko N, Demchenko P, Ślawska-Waniewska A, Dłużewski P, Greneche JM (2014) A magnetic nanogel based on O-carboxymethyl chitosan for antitumor drug delivery: synthesis, characterization and in vitro drug release. Soft Matter 10:34–41 21. Song Q, Zhang Z, Gao J, Ding C (2011) Synthesis and property studies of N-carboxymethyl chitosan. J Appl Polym Sci 119:3282–3328 22. Felicio SGF, Sierakowski MR, Petkowicz C, Silveira JLM (2008) Spherical aggregates obtained from N-carboxymethylation and acetylation of chitosan. Colloid Polym Sci 286: 1387–1394 23. Lonchamp J, Akintoy M, Cleg PS, Euston SR (2020) Functional fungal extracts from the Quorn fermentation co-product as novel partial egg white replacers. Eur Food Res Technol 246:69–80 24. Hayes ER (1986) N,O-carboxymethyl chitosan and preparative method therefor, US Patent, A4619995 25. Chen SC, Wu YC, Mi FL, Lin YH, Yu LC, Sung HW (2004) A novel pH-sensitive hydrogel composed of N,O-carboxymethyl chitosan and alginate cross-linked by genipin for protein drug delivery. J Control Release 96:285–300 26. Hassanabad AF, Zarzycki AN, Jeon K, Dundas JA, Vasanthan V, Deniset JF, Fedak PWM (2021) Prevention of post-operative adhesions: a comprehensive review of present and emerging strategies. Biomol Ther 11:1027 27. Anitha A, Sreeranganathan M, Chennazhi KP, Lakshmanan VK, Jayakumar R (2014) In vitro combinatorial anticancer effects of 5-fluorouracil and curcumin loaded N, O-carboxymethyl chitosan nanoparticles toward colon cancer and in vivo pharmacokinetic studies. Eur J Pharm Biopharm 88:238–251 28. Chen Y, Liu Y, Tan H, Jiang J (2009) Synthesis and characterization of a novel superabsorbent polymer of N,O-carboxymethyl chitosan graft copolymerized with vinyl monomers. Carbohydr Polym 75:287–292 29. Pandian M, Selvaprithviraj V, Pradeep A, Rangasamy J (2021) In-situ silver nanoparticles incorporated N, O-carboxymethyl chitosan based adhesive, self-healing, conductive, antibacterial and anti-biofilm hydrogel. Int J Biol Macromol 188:501–511 30. Anitha MS, Deepa N, Chennazhi KP, Nair SV, Jayakumar R (2012) Curcumin-loaded N, O-carboxymethyl chitosan nanoparticles for cancer drug delivery. J Biomater Sci Polym Ed 2:381–400 31. Wang L, Wang A (2008) Adsorption behaviors of Congo red on the N,O-carboxymethylchitosan/montmorillonite nanocomposite. J Chem Eng 143:43–50 32. Muzzarelli RAA, Ramos V, Stanic V, Dubini B, Mattioli-Belmonte M, Tosi G, Giardino R (1998) Osteogenesis promoted by calcium phosphate N,N-dicarboxymethyl chitosan. Carbohydr Polym 36:267–276 33. Mattioli-Belmonte M, Gigante A, Muzzarelli RAA, Politano R, De Benedittis A, Specchia N, Buffa A, Biagini G, Greco F (1999) N,N-dicarboxymethyl chitosan as delivery agent for bone morphogenetic protein in the repair of articular cartilage. Med Biol Eng Comput 37:130–134

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Adv Polym Sci (2024) 292: 31–44 https://doi.org/10.1007/12_2023_166 © The Author(s), under exclusive license to Springer Nature Switzerland AG 2023 Published online: 12 October 2023

Carboxymethyl Chitosan Derivatives and Its Interaction with Metal Ions Antonysamy Jeyaseelan, Natrayasamy Viswanathan, C. Sairam Sundaram, and S. Meenakshi

Contents 1 Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1.1 Chitosan . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1.2 Chitosan Derivatives . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1.3 Carboxymethyl Chitosan . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1.4 Heavy Metals and Its Toxicity . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2 Synthesis of Carboxymethyl Chitosan . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3 CMCS Interactions with Various Metal Ions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.1 CMCS Interaction with Divalent Metal Ions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.2 CMCS Interactions with Higher Valance Metal Ions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4 Conclusions and Perspective . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

32 32 32 33 33 35 36 36 38 39 41

Abstract Chitosan (CS) is a non-toxic, biocompatible, and biodegradable deacetylated derivative of chitin. CS was effective metal-chelator; however, its practical application is restricted because of adsorption capacities, stability in acid medium, and adsorption selectivity. Because of the existence of amine (–NH2) and hydroxyl (–OH) groups, CS could be altered to make different derivatives with varying solubility pattern. Chitosan chemical modification and its derivatives by graft copolymerization could improve their properties and as a result expand their

A. Jeyaseelan and N. Viswanathan Department of Chemistry, Anna University, University College of Engineering – Dindigul, Reddiyarchatram, Dindigul, Tamil Nadu, India C. Sairam Sundaram Department of Chemistry, Women’s Engineering College, Lawspet, Puducherry, India S. Meenakshi (✉) Department of Chemistry, The Gandhigram Rural Institute – Deemed to be University, Gandhigram, Dindigul, Tamil Nadu, India

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promising applications onto various fields. Carboxymethyl chitosan is one of the chitosan derivatives and it was most intensively explored because of its water solubility in broad pH conditions. However, it was employed in various promising applications. Carboxymethyl chitosan (CMCS) has soluble nature, hence it is difficult to use for metal ion removal studies. To overcome this problem, carboxymethyl chitosan was modified as insoluble hybrid material and was utilized for heavy metal removal studies by adsorption method. In this review, CMCS derivatives and its interaction with heavy metal ions were investigated in detail. It is hopefully predictable that the content will offer conclusive insights on the utilization of this biological activity, biodegradation, low-toxicity, and biocompatibility of CMCS and its interaction with various heavy metals for water purification and trust to boost up its promising application as heavy metals removal. Keywords Carboxymethyl · Chitosan · Adsorption · Chitosan · Metal ion removal

1 Introduction 1.1

Chitosan

Chitosan (CS) is one of the cationic copolymers from N-acetylglucosamine and glucosamine, which is a partly deacetylated derivative of chitin. In addition, CS is the majority abundant carbohydrate and also which is extracted from the exoskeleton of crustaceans [1]. CS has many unique features/characteristics like non-toxicity, biocompatibility, eco-friendliness, bio-adhesivity, biodegradability, and bio-renewability; these unique features and properties make it a polymer of reckoning [2]. Moreover, CS and its derivatives were employed in numerous promising applications like water treatment, pharmaceuticals, agriculture, food industry, biomedicine, cosmetics, etc. Though the promising established applications of CS suffer from stern drawbacks because of CS insolubility in alkaline/neutral pH medium and more stable crystalline network occurring from very strong hydrogen bonds, the most probable solubility was noted only in acidic medium under pH 6.5 (under the pKa value of CS) [3].

1.2

Chitosan Derivatives

The chitosan solubility could be modified with its chemical modifications and depolymerization. CS has active functional groups like amino and primary/secondary hydroxyl groups which could be employed for chemical functionalization/ modifications in mild reaction circumstances to modify its nature [4]. Water-soluble derivatives were developed by introducing hydrophilic groups (dihydroxyethyl,

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hydroxyalkylamino, hydroxypropyl); phosphate; sulfate; by grafting water-soluble polymers; quaternarization method; carboxyalkyl groups (carboxybutyl carboxymethyl, carboxyethyl) in chitosan macromolecular chain [5]. Compared to other water-soluble chitosan derivatives, CMCS has been promisingly and broadly investigated due to its simple synthesis, ampholytic behavior, and the potential applications of sample.

1.3

Carboxymethyl Chitosan

CMCS is one of the most significant derivatives from chitosan. Carboxymethyl chitosan is the product of carboxylation of chitosan holding carboxymethyl substituents on amino and primary hydroxyl active sites of the glucosamine units [6]. Carboxymethylation of chitosan is a new way for alteration of chitosan into a water-soluble form. CMCS has numerous unique physical, biological, and chemical properties like biocompatibility, low-toxic nature, and excellent capability to form hydrogels, fibers, and films [7]. Due to its non-toxicity, hydrophilicity, metalchelating ability, biodegradability, and eco-friendly properties, carboxymethyl chitosan stands out as excellent candidate among bio-adsorption materials. Therefore, CMCS has been potentially utilized in various fields like water purification, agriculture, food industry, and biomedicine [8]. As CMCS could not be employed to recover metal ions because of its water-soluble nature, modified hybrid materials of CMCS were developed to recover metal ions.

1.4

Heavy Metals and Its Toxicity

Heavy metals are basically defined as metallic chemical elements that have density of greater than 5 g/cm3. Heavy metals are released into the environment from different natural and anthropogenic resources. Natural sources are distributed in various ways like soil erosion, sedimentation, metamorphic rocks, volcanic activity, and forest fires [9]. The heavy metal ores like granite (Mn & Cr), hematite (Fe), sphalerite (Zn), molybdenite (Mo), gold veins in rock (Au) and chalcopyrite (Cu) are mostly occupied in the soil. Heavy metals are released into the environment by man-made activities like industries, agricultural activities, and sewage effluents. In addition, the atmospheric heavy metals like lead, cadmium, etc. also enter into the aquatic ecosystem by aerosols, gases, and particulates. Atmospheric heavy metal sources are volcanic aerosols, forest fires, mineral dusts, sea salt particles, coal combustions, transportation emissions, and atomic radiations. The anthropogenic sources of heavy metals along with the tolerance limit according to WHO and their toxicity are listed in Table 1. Aqueous effluents from various sources like tannery, pigment, textile, electroplating, paint, metallurgical industries majorly contain heavy metal

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Table 1 Sources of heavy metals with tolerance limit and their effects Heavy metals Arsenic (As) Cadmium (Cd) Chromium (Cr) Lead (Pb)

Anthropogenic sources Algaecides, herbicides, insecticides, fungicides, smelting operations, and metallurgical industries Metal plating, pesticides industries, and pigments Leather industries, mining, tannery industries, and metal finishing Pesticide industries, lead acid battery and paints

Tolerance limit (mg/g) 0.01

0.003 0.05 0.01

Mercury (Hg)

Amalgamation, electronics, and coal combustion industries

0.006

Iron (Fe)

Metallurgical, steel manufacturing, food and medicine industries

1.0

Nickel (Ni)

Battery industries and thermal power plants

0.05

Zinc (Zn)

Pyrometallurgical, cosmetics, soaps, and batteries

0.5

Health effects Kidney failure, keratosis, skin cancer, visceral cancer Phytotoxic, itai-itai disease and kidney failure Carcinogenic, kidney and liver damage Effects to brain, nervous, circulatory, and kidney function Biomagnification in aquatic environments, Minamata disease Genetic disorder, stomach problems, nausea and vomiting Contact dermatitis, asthma, chronic respiratory infections and carcinogenic Hypertension, lethargy, neurotic effects and affects cholesterol metabolism

pollutants. The major heavy metal pollutants present in effluents were nickel, copper, cadmium, mercury, arsenic, chromium, zinc, lead, etc. The heavy metal removal from industrial effluents and wastewater is a vital challenge for researchers due to its very low concentration as well as cost and effort. These metals create significant risks to the environment at low concentration because they are non-biodegradable, carcinogenic and have tendency to accumulate with human organs which leads to severe diseases [10]. They not only generate the severe water pollution and also affect the other water quality parameters. Hence, the removal of heavy metal from water is a necessary one. Nowadays various conventional techniques like chemical precipitation, electrochemical techniques, membrane separation, coagulation & sedimentation, ion exchange, biological method, and adsorption are used [11]. Among them adsorption method was more efficient, low cost, simple, high efficacy, and easy technique. Different adsorption materials like metal oxides, clay, biopolymer, activated carbon, and polyvinyl alcohol based materials mostly employed for heavy metal adsorption. However, these materials have numerous limitations like poor adsorption capacity/selectivity. To enhance the adsorption capacity, the CMCS and its derivatives-based hybrid material were developed and it was employed for heavy metal removal. Therefore, this review is intended to highlight the carboxymethyl chitosan derivatives and its interaction with heavy metal ions from water. Carboxymethyl chitosan

Carboxymethyl Chitosan Derivatives and Its Interaction with Metal Ions

35

Fig. 1 Graphical representation of the applications, composite types, and adsorption studies of CMCS

derivatives were widely utilized in water purification because of its biocompatibility, high water stability, metal-chelating ability, biodegradability, and eco-friendly. The comprehensive study about carboxymethyl chitosan derivatives with their interaction with heavy metal ion is focused in detail. The graphical representation of the applications, composite types, and adsorption studies of CMCS was depicted in Fig. 1.

2 Synthesis of Carboxymethyl Chitosan Carboxymethyl chitosan was developed using chloroacetate and alcoholic solvent in isopropyl alcohol and water mixture [12]. Initially, about 10 g of polysaccharide was entirely suspended in 10 mL of isopropyl alcohol then 26 mL of NaOH (10 mol L-1) solvent was subsequently added into the above mixture and the mixture of content was mixed well for 45 min. The reaction was initiated by the addition of 30 g of

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chloroacetic acid and performed with continuous stirring for 2 h at 60°C. Then, the reaction was stopped by addition of deionized water (10 mL) and neutralized with concentrated acetic acid (10 mL). The mixture was dialyzed and lyophilized.

3 CMCS Interactions with Various Metal Ions 3.1 3.1.1

CMCS Interaction with Divalent Metal Ions CMCS Interactions with Ni(II) and Cu(II)

Kavitha et al. developed the water-soluble chitosan derivatives like N-N-Ntriethylammonium chitosan (TEAC) and carboxymethyl chitosan (CMCh) which were prepared and utilized for Cu(II), Ni(II), and Cr(VI) removal by size enhanced ultrafiltration. CMCh consist of –NH2 and –COOH moieties, which make possible attractions with Cu(II) and Ni(II) via complexation process. CMCh has -NH2 and carboxymethyl (-CH2-COOH) groups in its structure. Cu(II) and Ni(II) are expected to complex with -CH2-COO-. The proposed mechanism of CMCh and TEAC for Ni (II), Cr(VI), and Cu(II) removal involved was complexation and decomplexation [13]. Sun et al. investigated the N,O-carboxymethyl-chitosan (N,O-CMC) for effective Cu(II) adsorption. The high adsorption capacity of N,O-CMC for Cu(II) was found to be 162.5 mg/g. During adsorption, the chemical bond and chelation were formed between the N,O-CMC and Cu(II) ions [14]. He et al. prepared carboxymethyl chitosan (CMCS)-Kaolin hydrogel via simple Schiff base reaction for effective copper ion removal. The Cu2+ adsorption capacity of CMCS-Kaolin hydrogels was 206 mg/g. The Cu2+ removal by CMCS-Kaolin hydrogels was done by ion exchange of protonated –NH2 and chelation adsorption of –COOH [15]. Zhang et al. fabricated magnetic bentonite/carboxymethyl chitosan/sodium alginate (Mag-Ben/CCS/Alg) for Cu(II) adsorption. The fabricated Mag-Ben/CCS/Alg has achieved Cu(II) adsorption capacity of 56.79 mg/g and percent removal was 92.62% within 90 min at 30°C. Adsorption kinetic results exhibit that the mechanism of Cu(II) on Mag-Ben/CCS/Alg followed chemical adsorption and ion exchange process [16]. Weerasinghe et al. developed the bi-functional chitosan derivative, like ethylenediaminetetraacetic acid–carboxymethyl chitosan (EDTACMC) with connection of carboxymethyl and EDTA on polymer backbone and so raising its metal-binding nature. The Cu2+ uptake capacity of EDTA-CMC was 111.90 mg/g using 10 mg/L initial Cu2+ solution [17]. Zheng et al. developed carboxymethyl chitosan (CMCS) using adipic acid dihydrazide as cross-linker for Cu2+ removal. The highest adsorption capacity of Cu2+ was 200 mg/g using CMCS within 10 min. The major mechanism of Cu2+ on CMCS was ion exchange and surface complexation [18]. Shehzad et al. developed alginate/carboxymethyl chitosan/TiO2 (TiO2/TSC-CMC) [amino-thiocarbamate derivative] for Ni (II) removal from water. The highest adsorption capacity of TiO2/TSC-CMC composite was 172 mg/g at pH 6. Ni(II) adsorption on TiO2/

Carboxymethyl Chitosan Derivatives and Its Interaction with Metal Ions

37

TSC-CMC occurs mainly via ion exchange and inner complexation mechanism [19]. Tian et al. developed the polyacrylic acid (PAA) with CMCS (PAA-CMCS) via grafting copolymerization technique for Ni(II) removal from wastewater. The highest Ni(II) adsorption capacity of PAA-CMCS was 200.588 mg/g and major mechanism of Ni(II) on PAA-CMCS was surface adsorption, intraparticle diffusion, chelation, and coordination [20].

3.1.2

CMCS Interactions with Zn(II), Pb(II), Co(II), Sr(II), and Hg(II)

Song et al. prepared xanthated carboxymethyl chitosan (XCC) from xanthation reaction of N-carboxymethyl chitosan (NCMC) for Pb(II) adsorption. The highest adsorption capacity of XCC for Pb(II) adsorption was 520.8 mg/g. XPS and FTIR results revealed that nitrogen, carboxyl groups, and sulfur groups of XCC were involved in the surface interactions and complexation process with Pb(II) adsorption [21]. Zhu et al. developed ZIF-8 incorporated carboxymethyl chitosan beads (BCMC), namely BCMC@ZIF-8 for Pb2+ removal. XPS and kinetic results exhibit the mechanisms like diffusion and sharing/transfer of electrons between Pb2+ and BCMC@ZIF-8. The Pb2+ adsorption capacity of 566.09 mg/g using BCMC@ZIF8 was from Langmuir model [22]. Wang et al. developed polyethylenimine (PEI) modified magnetic carboxymethyl chitosan (MCMC-PEI) for Pb(II) removal from water. The enhanced Pb (II) adsorption capacity of MCMC-PEI was 124.0 mg/g from Langmuir-Freundlich model. The surface complexation and/or chemical reaction was the major Pb (II) removal mechanism [23]. Kavitha et al. developed the biodegradable and water-soluble carboxymethyl chitosan (CMCh) and employed for heavy metals Pb (II), Zn(II), Cu(II), and Ni(II) separation from water which heavy metals are the main industrial contaminants present in wastewater. The highest Pb(II), Zn(II), Cu(II), and Ni(II) rejection percentage of CMCh was 100, 100, 95, and 98%, respectively, at pH -10 with initial Pb(II), Zn(II), Cu(II), and Ni(II) concentration, respectively, at 70 mg/L. The CMCh-metal ions complex was investigated via morphology and elemental composition for CMCh and CMCh complex with metal ions Pb(II), Zn(II), Cu(II), and Ni(II). The EDAX results prove the complexation of CMCh with metal ions like Pb(II), Zn(II), Cu(II), and Ni(II). The significant peaks like C, O, and N related to CMCh were complexed with corresponding Pb(II), Zn(II), Cu(II), and Ni (II) ions. In pH studies, with pH from 4 to 7, the Cu(II) removal percentage raises from initial pH and after pH 7 there is no significant Cu(II) removal with CMCh. With increase raises in initial pH from 7, the Cu(II) removal percentage was >90% and it was moderately less in acidic pH condition. The raises in the Cu (II) removal percentage are mainly due to carboxyl moiety existing in CMCh which competes with OH- for complexation with Cu(II), however at acidic pH medium, H+ ions battle with polymer ligands for complexation with Cu(II). This action is only noted until pH 7. After pH 7 to 10, there is no change in Cu(II) removal percentage though pH 7 is a favorable condition for complexation process, because of the excess Cu(II) availability for complexation with CMCh [24].

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Charpentier et al. developed magnetic carboxymethyl chitosan (CMC) nanoparticles by simple one-step chemical coprecipitation technique for Cu2+, Zn2+, and Pb2+ and ions removal from water. The maximum uptakes of Pb(II) is 107.5 mg/g, Cu(II) is 99 mg/g, and Zn(II) is 75.8 mg/g using CMC composite, respectively. The enhanced Pb2+, Cu2+, and Zn2+ adsorption capacities is due to accessibility of bulky -COO- groups which has the ability to coordinate with Pb2+, Cu2+, and Zn2+ metal ions by chelation coordination as major mechanism for Pb2+, Cu2+, and Zn2+ removal [25]. Carboxymethyl chitosan-g-polymethylmethacrylate (CMCS-g-PMMA) was developed by microwave-assisted route and it was utilized for Hg2+ metal ion removal. The enhanced Hg2+ adsorption capacity was 87.71 mg/g within 90 min from Langmuir isotherm. The Hg2+ on CMCS-g-PMMA was followed intermolecular forces attraction and physical adsorption (van der Waals (weak intermolecular) interactions) between adsorbent and adsorbate [26]. Luo et al. developed the porous carboxymethyl chitosan (PCMC) beads by coacervation/chemical cross-linking by polyethylene glycol and CaCl2 and glutaraldehyde as cross-linkers for Co(II) removal. The PCMC beads have the Co (II) adsorption capacity of 46.25 mg/g within 6 h. The Co(II) interacts with PCMC beads by chemical adsorption [27]. Mei et al. developed the carboxymethyl chitosan (O-CMC) with functionalizing and cross-linking by ethylenediaminetetraacetic acid via simple one-pot synthesis under mild conditions. The maximum Sr(II) adsorption capacity of O-CMC/EDTA was 105.81 mg/g within neutral pH. The mechanism of removal of Sr (II) by O-CMC followed complexation and hydrogen bond [28]. The comprehensive summary of adsorption mechanism of CMCS toward metal ions was given in Fig. 2.

3.2

CMCS Interactions with Higher Valance Metal Ions

Borsagli et al. developed the carboxymethyl chitosan (CMC) and employed for Cr (VI) removal. The developed CMC adsorbed Cr(VI) via complexation process. The efficient carboxylic groups incorporation in chitosan (CMC) drastically modified the complexation and adsorption mechanism for Cr(VI) removal. The Cr(VI) on CMC was majorly followed complexation [29]. Wu et al. developed the carboxymethyl chitosan–hemicellulose resin (CMCH) with thermal cross-linking method for Cr (VI) and Mn(VII) removal from water. The greatest adsorption capacity of CMCH for Mn(VII) and Cr(VI) was 42.0 and 49.0 mg/g, respectively. The CMCH with metal ions (Mn(VII) and Cr(VI)) formed complexation process and this was verified with FTIR analysis [30]. The ternary composite, namely carboxymethyl chitosan, hemicellulose, and nanosized TiO2 (CHNT) prepared by TiO2 nanoparticles incorporated with carboxymethyl chitosan-hemicellulose. The developed CHNT was investigated for toxic heavy metals (Mn(VII) and Cr(VI)) removal. The higher removal capacity was observed because of its preferable chelating groups in structure with metal ions [31]. Deng et al. developed carboxymethyl chitosan-based membrane using

Carboxymethyl Chitosan Derivatives and Its Interaction with Metal Ions

39

Fig. 2 The general adsorption mechanism of CMCS and its derivatives toward metal ions

carboxymethyl chitosan and silicon dioxide for toxic Cr(VI) removal. The high Cr (VI) removal of membrane was 80% at 60 min with pH 5 and 40 mg/L initial Cr (VI) concentration. The carboxymethyl chitosan-based membrane has free amino groups and hydroxyl groups and these interact with Cr(VI) by chelation mechanism [32]. It could be observed that carboxymethyl chitosan effectively removes lower valence ions than higher valence heavy metal ions. The adsorption capacity comparison of CMCS derivatives for various metal ions was given in Table 2.

4 Conclusions and Perspective Water is a natural resource essential for the survival of all the living things. Pure and unpolluted water is required for equality of environment in the world. Water pollutants are stimulated too many diseases on humans, animals, and aquatic organisms. Among them, water contamination from the toxic metals has gained significant attention because of their harmful effects. Heavy metals are released from various sources from the production of dyes, papers, chrome plating, paints, pharmaceutical and leather industries. Excess heavy metal containing water causes various disorders such as carcinogenic effect, mutagenic effect, kidney damage, and liver damage on human beings. Hence, WHO has fixed a tolerance limit of heavy metal concentration in potable water.

40

Table 2 Adsorption capacity comparison of CMCS derivatives for metal ions Adsorption capacity (mg/g) S. no. 1 2 3 4 5 6 7 8 9

Cu (II) 162.50 206.00 56.79 111.90 – – – – –

Ni (II) – – – – 172.19 250.58 – – –

Pb (II) – – – – – – 520.80 566.09 124.00

Zn (II) – – – – – – – – –

Co (II) – – – – – – – – –

Sr (II) – – – – – – – – –

Hg (II) – – – – – – – – –

Cr (VI) – – – – – – – – –

Mn (VII) – – – – – – – – –

References [13] [15] [16] [17] [19] [20] [21] [22] [23]

99.00 – – – –

– – – – –

107.50 – – – –

75.80 – – – –

– – 46.25 – –

– – – 105.81 –

– 87.71 – – –

– – – – 49.00

– – – – 42.00

[25] [26] [27] [28] [30]

A. Jeyaseelan et al.

10 11 12 13 14

Adsorbents name N,O-CMCS CMCS-Kaolin hydrogel Magnetic bentonite/CMCS/sodium alginate Ethylenediaminetetraacetic acid–CMCS Alginate/carboxymethyl chitosan/TiO2 Polyacrylic acid functionalized CMCS Xanthated CMCS ZIF-8 incorporated CMCS beads Polyethylenimine enhanced magnetic CMCS Magnetic CMCS material CMCS-g-polymethylmethacrylate Porous CMCS O-CMCS CMCS–hemicellulose resin

Carboxymethyl Chitosan Derivatives and Its Interaction with Metal Ions

41

Numerous methods have been developed for the heavy metal removal from water. Among the existing techniques, the adsorption seems to be a fascinating method to remove heavy metal from water. Because adsorption process offers many features such as economic feasibility, flexibility in design, and high selectivity, adsorbents can be regenerated by suitable eluent. Plenty of materials have been used as adsorbent for the heavy metal removal from water which includes inorganic materials, polymeric materials, carbon-based materials, natural minerals, organic materials, etc. Biopolymers like chitosan, chitin, cellulose, and sodium alginate have found wide range of applications in removing trace metals from aqueous solution due to its biodegradability, biocompatibility, and non-toxicity. CMCS is one of the most significant derivatives from chitosan. Carboxymethyl chitosan is the product of carboxylation of chitosan holding carboxymethyl substituents on amino and primary hydroxyl active sites of glucosamine units. Carboxymethyl chitosanderived materials were developed toward heavy metal ion interactions. This review summarizes research on carboxymethyl chitosan with potential applications in metal ion removal in water treatment. The properties of chitosan and its derivatives could be modified by chemical modification. Carboxymethyl chitosan derivatives and its interaction with various heavy metal ions were studied and it was utilized for treatment of wastewater. The removal efficiency of CMC and modified CMCs decreases when we move on to higher valent heavy metal ions. It is hopefully predictable that the content will offer conclusive insights on the utilization of this biological activity, biodegradation, low-toxicity, and biocompatibility of carboxymethyl chitosan, its interaction with various heavy metals for water purification, and trust to boost up its promising application as heavy metal removal.

References 1. El-Sherbiny IM (2009) Synthesis, characterization and metal uptake capacity of a new carboxymethyl chitosan derivative. Eur Polym J 45:199–210 2. Kausar A (2017) Scientific potential of chitosan blending with different polymeric materials: a review. J Plast Film Sheeting 33:384–412 3. Ryu JH, Lee Y, Kong WH, Kim TG, Park TG, Lee H (2011) Catechol-functionalized chitosan/ pluronic hydrogels for tissue adhesives and hemostatic materials. Biomacromolecules 12:2653– 2659 4. Wang W, Meng Q, Li Q, Liu J, Zhou M, Jin Z, Zhao K (2020) Chitosan derivatives and their application in biomedicine. Int J Mol Sci 21:487 5. Jabbal-Gill I, Watts P, Smith A (2012) Chitosan-based delivery systems for mucosal vaccines. Expert Opin Drug Deliv 9:1051–1067 6. Tzaneva D, Simitchiev A, Petkova N, Nenov V, Stoyanova A, Denev P (2017) Synthesis of carboxymethyl chitosan and its rheological behaviour in pharmaceutical and cosmetic emulsions. J Appl Pharm Sci 7:070–078 7. Mourya VK, Inamdara N, Ashutosh Tiwari N (2010) Carboxymethyl chitosan and its applications. Adv Mater Let 1:11–33 8. Zargar V, Asghari M, Dashti A (2015) A review on chitin and chitosan polymers: structure, chemistry, solubility, derivatives, and applications. Chem Bio Eng Rev 2:204–226

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9. Sardar K, Ali S, Hameed S, Afzal S, Fatima S, Shakoor MB, Bharwana SA, Tauqeer HM (2013) Heavy metals contamination and what are the impacts on living organisms. Greener J Environ Manage Public Safety 2:172–179 10. Isangedighi IA, David GS (2019) Heavy metals contamination in fish: effects on human health. J Aquatic Sci Mar Biol 2:7–12 11. Azimi A, Azari A, Rezakazemi M, Ansarpour M (2017) Removal of heavy metals from industrial wastewaters: a review. Chem Bio Eng Rev 4:37–59 12. Taubner T, Marounek M, Synytsya A (2020) Preparation and characterization of hydrophobic and hydrophilic amidated derivatives of carboxymethyl chitosan and carboxymethyl β-glucan. Int J Biol Macromol 163:1433–1443 13. Kavitha E, Sowmya A, Prabhakar S, Jain P, Surya R, Rajesh MP (2019) Removal and recovery of heavy metals through size enhanced ultrafiltration using chitosan derivatives and optimization with response surface modeling. Int J Biol Macromol 132:278–288 14. Sun S, Wang A (2006) Adsorption kinetics of Cu (II) ions using N, O-carboxymethyl-chitosan. J Hazard Mater 131:103–111 15. He G, Wang C, Cao J, Fan L, Zhao S, Chai Y (2019) Carboxymethyl chitosan-kaolinite composite hydrogel for efficient copper ions trapping. J Environ Chem Eng 7:102953 16. Zhang H, Omer AM, Hu Z, Yang LY, Ji C, Ouyang XK (2019) Fabrication of magnetic bentonite/carboxymethyl chitosan/sodium alginate hydrogel beads for Cu (II) adsorption. Int J Biol Macromol 135:490–500 17. Weerasinghe K, Liyanage S, Kumarasinghe UR, Cooray AT (2022) Synthesis of a bifunctional EDTA–carboxymethyl chitosan derivative and its potential as an adsorbent for the removal of Cu2+ ions from aqueous solutions. Polym Renew Resour 13:170–187 18. Zheng E, Dang Q, Liu C, Fan B, Yan J, Yu Z, Zhang H (2016) Preparation and evaluation of adipic acid dihydrazide cross-linked carboxymethyl chitosan microspheres for copper ion adsorption. Colloids Surf A Physicochem Eng Asp 502:34–43 19. Shehzad H, Farooqi ZH, Ahmed E, Sharif A, Din MI, Arshad M, Nisar J, Zhou L, Yun W, Nawaz I, Hadayat M (2020) Fabrication of a novel hybrid biocomposite based on aminothiocarbamate derivative of alginate/carboxymethyl chitosan/TiO2 for Ni (II) recovery. Int J Biol Macromol 152:380–392 20. Tian T, Bai Z, Wang B, Zhao S, Zhang Y (2020) Facile fabrication of polyacrylic acid functionalized carboxymethyl chitosan microspheres for selective and efficient removal of Ni (II) from multicomponent wastewater. Colloids Surf A Physicochem Eng Asp 597:124676 21. Song Q, Wang C, Zhang Z, Gao J (2014) Adsorption of lead using a novel xanthated carboxymethyl chitosan. Water Sci Technol 69:298–304 22. Zhu X, Tong J, Zhu L, Pan D (2022) In situ growth of ZIF-8 on carboxymethyl chitosan beads for improved adsorption of lead ion from aqueous solutions. Int J Biol Macromol 205:473–482 23. Wang Y, Wu D, Wei Q, Wei D, Yan T, Yan L, Hu L, Du B (2017) Rapid removal of Pb(II) from aqueous solution using branched polyethylenimine enhanced magnetic carboxymethyl chitosan optimized with response surface methodology. Sci Rep 7:10264 24. Kavitha E, Kedia R, Babaria N, Prabhakar S, Rajesh MP (2020) Optimization of process using carboxymethyl chitosan for the removal of mixed heavy metals from aqueous streams. Int J Biol Macromol 149:404–416 25. Charpentier TV, Neville A, Lanigan JL, Barker R, Smith MJ, Richardson T (2016) Preparation of magnetic carboxymethylchitosan nanoparticles for adsorption of heavy metal ions. ACS Omega 1:77–83 26. Labidi A, Salaberria AM, Labidi J, Abderrabba M (2019) Preparation of novel carboxymethylchitosan-graft-poly (methylmethacrylate) under microwave irradiation as a chitosan-based material for Hg2+ removal. Microchem J 148:531–540 27. Luo W, Bai Z, Zhu Y (2018) Fast removal of Co (II) from aqueous solution using porous carboxymethyl chitosan beads and its adsorption mechanism. RSC Adv 8:13370–13387 28. Mei J, Mo S, Zhang H, Zheng X, Li Z (2020) Removal of Sr(II) from water with highly-elastic carboxymethyl chitosan gel. Int J Biol Macromol 163:1097–1105

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29. Borsagli FGM, Mansur AA, Chagas P, Oliveira LC, Mansur HS (2015) O-carboxymethyl functionalization of chitosan: complexation and adsorption of Cd (II) and Cr (VI) as heavy metal pollutant ions. React Funct Polym 97:37–47 30. Wu SP, Dai XZ, Kan JR, Shilong FD, Zhu MY (2017) Fabrication of carboxymethyl chitosan– hemicellulose resin for adsorptive removal of heavy metals from wastewater. Chin Chem Lett 28:625–632 31. Wu S, Kan J, Dai X, Shen X, Zhang K, Zhu M (2017) Ternary carboxymethyl chitosanhemicellulose-nanosized TiO2 composite as effective adsorbent for removal of heavy metal contaminants from water. Fibers Polym 18:22–32 32. Deng Y, Kano N, Imaizumi H (2017) Adsorption of Cr(VI) onto hybrid membrane of carboxymethyl chitosan and silicon dioxide. J Chem 2017:3426923

Adv Polym Sci (2024) 292: 45–58 https://doi.org/10.1007/12_2023_162 © The Author(s), under exclusive license to Springer Nature Switzerland AG 2023 Published online: 21 September 2023

Antimicrobial Properties of Carboxymethyl Chitosan Derivatives and Its Composites Vidyaalakshmi Venkatesan and R. Jayakumar

Contents 1 Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2 Antibacterial Activity of CMC . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3 Antifungal Activity of CMC . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4 Conclusion . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

45 48 52 54 54

Abstract Chitosan and its derivatives are being widely used in the biomedical field mainly because of its properties like biodegradability, biocompatibility, and antimicrobial activity. The main disadvantage of parent chitosan is the poor water solubility. The water-soluble, carboxymethyl chitosan derivative has better properties leading it to be a more suitable choice in cases of drug delivery, tissue engineering, hemostasis, and wound healing. Carboxymethyl chitosan derivative had shown significant antibacterial and antifungal activity which have been studied. This review mainly discusses the antibacterial and antifungal activity of carboxymethyl chitosan in detail. Keywords Antibacterial · Antifungal · Carboxymethyl chitosan · Composites · Microbial infections

V. Venkatesan and R. Jayakumar (✉) Polymeric Biomaterials Lab, School of Nanosciences and Molecular Medicine, Amrita Vishwa Vidyapeetham, Kochi, India e-mail: [email protected]

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1 Introduction Chitosan, β-(1,4)-linked polysaccharide of D-glucosamine, is a biopolymer synthesized by the deacetylation of chitin. Chitosan is a water-insoluble cationic polymer, and the charge density depends on the degree of acetylation and the pH of the media. The derivatives of chitosan are made in order to improve the inherent properties of chitosan such as biodegradability and solubility as well as to incorporate new properties and functions [1]. One of the important water-soluble chitosan derivatives is carboxymethyl chitosan (CMC). CMC is prepared by substituting –OH or –NH2 groups of chitosan with –CH2COOH groups. The important carboxymethyl chitosan types are N, O-carboxymethyl chitosan, O-carboxymethyl chitosan, N-carboxymethyl chitosan, N,N-dicarboxymethyl chitosan, O-succinyl chitosan, N-succinyl chitosan, N-carboxybutyl chitosan, and N-carboxyethyl chitosan and are shown in Fig. 1 [2]. CMC has unique physical, chemical, and biological properties like high viscosity, hydrodynamic volume, low toxicity, biocompatibility, and water solubility. It is being widely used in biomedical fields as a bactericide, wound dressing/moistureretention agent, artificial skin and bone, drug delivery, and hemostasis [3]. CMC

Fig. 1 Different types of carboxymethyl chitosan derivatives

Antimicrobial Properties of Carboxymethyl Chitosan Derivatives and. . .

47

Fig. 2 Antibacterial mechanistic action of CMC

Fig. 3 Mechanism of fungal cell lysis by the interaction of the CMC with the fungal cell membrane

inhibits the bacterial growth by aggregation through charge neutralization and flocculation through bridging mechanism [4]. CMC also hinders the conversion of DNA into RNA and thereby inhibiting the protein synthesis which is vital for the bacterial growth as seen in Fig. 2. In case of CMC antifungal property, it binds to the cell wall function causing inhibition, metabolic disturbances, and cellular reproduction and eventually leads to the death of the cell as seen in Fig. 3. CMC has a better antimicrobial activity as compared to that of chitosan due to the substitution of the alkyl chains on the chitosan molecule. Weak NH3+ repulsion is caused due to the low NH2 protonation. The hydrophobic areas are exposed due to the formation of the intermolecular hydrogen bonds on the polymer chains leading to hydrophilic and hydrophobic areas, increasing the affinity between the wall of the cell and the chitosan derivative. This review entails the antibacterial and antifungal activity of the CMC derivatives and their composites in detail based on the reported studies.

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2 Antibacterial Activity of CMC The antibacterial activity increases from O-CMC to chitosan to N, O-CMC, the reason being the dependence of polycation antibacterial activity on the effective number of -NH3+ groups [5]. The silver nanoparticles were linked along with OCMC onto cotton surface fabric by a process known as mist modification. O-CMC binder was linked covalently to that cotton fabric by the process of esterification. The silver nanoparticles adhered tightly to the surface of the fiber with the help of coordination bonds with the amine groups of O-CMC. The incorporation of silver nanoparticles on the cotton fabric exhibited antibacterial properties against S. aureus and E. coli. It was also durable in a way that even after 50 cycles of washing the silver nanoparticles remained on the cotton fabric surface [6]. The hydrogel film produced from the O-CMC loaded waterborne polyurethane-gelatin hydrolysate exhibited antibacterial activity against E. coli and S. aureus. The antibacterial activity was increased as the pH value was lower with the higher concentration of H+ ions in the solution as there is more protonation of amino groups. The normal skin pH ranges between 4.2 and 5.6 where the outer layer of the skin which is acidic is vital for functioning of the skin and the inhibition of microbes, also wound healing occurs at an acidic pH. The prepared hydrogel was ideal for dressings of the wound area because of the high mechanical strength and also the antibacterial effect [7]. The supramolecular hydrogel using self-healing O-CMC cross-linked with zinc ions was developed. The supramolecular hydrogel system was developed by the metal ions solution added to the O-CMC at a suitable pH range. With the higher amount of zinc ions, the higher the antibacterial activity against S. aureus and E. coli. The antibacterial effect was hypothesized to be through the contact killing or release of zinc ions from the hydrogel. The positively charged hydrogel system interacts with the negatively charged cell wall. The zinc ions released from the hydrogel which are also positively charged could adhere to the negative cell wall through the electrostatic forces causing the cell wall damage. The zinc ions also bind to the nucleic acids and proteins of bacteria leading to the cellular distortion and cell death of the bacteria [8]. Hydrogel has a drawback of wear and tear at the wound site and to mitigate this issue, self-healing hydrogel made of N, O-CMC, and EDTA-ferric ion complex (EDTA: Fe3+) incorporated with the silver nanoparticles was fabricated. The prepared self-healing hydrogel possesses adhesiveness, injectability, and conductivity, also exhibited antibacterial activity against S. aureus, E. coli, K. pneumonia, P. aeruginosa, and MRSA. The reason of antibacterial activity is the binding of silver nanoparticles to the cell membrane which reduces its permeability and respiration [9]. Multifunctional blended membranes made of poly(vinyl alcohol) and OCMC are produced by a simple solution casting method for the application of wound dressings. The antibacterial effect was evident against S. aureus and E. coli as the content with chitosan was increased [10]. The Pickering emulsion strategy of electrospinning was used in wound dressings where there was a need to improvise citral loading in a hydrophilic nanofibrous material. The O-CMC and poly(vinyl

Antimicrobial Properties of Carboxymethyl Chitosan Derivatives and. . .

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Fig. 4 Site of action of O-CMC against Gram-positive cell membrane

alcohol) (PVA) and citral mixed solutions were made as the inner oil phase covered by the outer water phase is continuous and the cyclodextrin-citral inclusion complex particles act as emulsion stabilizers. The citral is believed to penetrate the lipid structure of the bacterial cell wall thereby causing proteins along with the cell membrane damage followed by cytoplasm leakage causing cell lysis and cell death [11]. An antibacterial adhesive was created by the addition of O-CMC and the polymerized chitosan significantly was able to decrease the S. mutans and the inhibition was comparatively more when the concentration of O-CMC was increased [12]. Antibacterial efficacy of O-CMC exhibited activity against Gram-negative bacteria like Enterococcus faecalis, Shigella, Flexneri, Klebsiella pneumoniae, Pseudomonas aeruginosa, Vibrio parahaemolyticus and the Gram-positive bacteria like Bacillus subtilis, Staphylococcus aureus, Bacillus cereus [13]. In case of Grampositive bacteria, the positively charged O-CMC electrostatically interacts with the negatively charged teichoic acids in the peptidoglycan layer, in turn causing the cell membrane destruction. The intracellular components leakage followed by the entry of the O-CMC into the cell is shown in Fig. 4. O-CMC cross-linked polyacrylate films exhibited antibacterial activity against E. coli and S. aureus where the O-CMC is known to have excess of -COOH. This could improve the antibacterial effect against both Gram-positive and negative bacteria. Alongside this, the increase in the content of O-CMC is known to have increased antibacterial activity. The S. aureus was more strongly inhibited than the E. coli because the double outer membrane formed by the E. coli is less prone to surface damage as compared to S. aureus which has got a single-cell membrane layer [14]. The nisin loaded O-CMC nanogel prepared by the combination method of

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electrostatically self-assembling and chemical cross-linking had shown excellent antibacterial activity against S. aureus and E. coli. The inhibition of the bacteria by the nisin is due to the cell wall formation suppression. The attachment of nisin to lipid II, which is a peptidoglycan precursor, prevents its synthesis [15]. A proposal modifying the guar gum through quaternization graft reaction and then the process of oxidation yields oxidized quaternized ammonium groups become the components of the Schiff-based hydrogel due to the aldehyde group present. Addition of this to CMC yields a hydrogel that had good antibacterial, self-repairing, hemostasis, as well as injectable properties. The main reason for showing antibacterial activity was due to the presence of a positive charge of the quaternary ammonium salt that interacts with the negative charge lipids, proteins, and the carbohydrate residues on the bacterial cell surface. This eventually leads to the bacterial inactivation and cell damage [16]. O-CMC-zinc and α-calcium sulfate hemihydrate prepared had proved to evoke the antibacterial properties due to release of the zinc ions and activity was seen against S. aureus and E. coli [17]. By incorporating cerium ions to cross-link the CMC with that of the sodium alginate had proven antibacterial activity due to the gradual release of the cerium ions from the combinatorial sphere against the S. aureus and E. coli [18]. The essential functions of surface of the intravascular catheters include the antimicrobial activity along with the antithrombotic and even the low-friction functions. The nano and micro porous O-CMC were fabricated using the selective elimination of water-soluble polyethylene glycol. This compound showed a significant hydration rate and super hydrophilic property. The antifouling nature of this super hydrophilic surface had shown anti-adhesion of bacteria E. coli. The benefits of super hydrophilic surface had proved to inhibit the formation of biofilm by the mechanism of anti-adhesion [19]. The immediate elimination of the bacteria from the wound site and excess free radicals’ elimination along with the wound immune microenvironment modulation favor in terms of enhanced wound healing rate. A cross-linked hydrogel prepared using sulfhydryl O-CMC with maleimide-based oxidized sodium alginate through click chemistry had proven to control the bacterial growth of E.coli and S. aureus [20]. The complications of titanium implantation are bacterial infection and loosening that leads to the reduction of the survival rate of the implant. A quaternary ammonium O-CMC, hydroxyapatite, and collagen multilayers coated on top of Ti substrate by the method of layer-bylayer technique exhibited antibacterial property against S. aureus by contact killing [21]. The pin tract infection is a major problem in the orthopedic setup and the implant coated with O-CMC along with zinc ion was found to be more efficacious against S. aureus infection [22]. The antibacterial activity of the viscose rayon/OCMC fibers developed by the spinning of mixture of O-CMC xanthate and the cellulose xanthate by the viscose process was shown against E. coli [23]. In case of Gram-negative bacteria, the lipopolysaccharides have high negative charges which can be neutralized by the positive charges of the O-CMC leading to the disruption of the outer membrane and thereby allowing the chitosan to penetrate deep into the bacterial cell as depicted in Fig. 5.

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Fig. 5 Site of action of O-CMC against Gram-negative cell membrane

The infection at the burn site was successfully treated using silver nanoparticles loaded silk fibroin/O-CMC which prevented S. aureus and P. aeruginosa which are particularly known for their drug resistance [24]. An injectable hydrogel based on OCMC and oligomeric procyanidin was developed, where the oligomeric procyanidin serves as a dynamic cross-linking agent for bridging O-CMC macromolecules essentially through the dynamic hydrogen bonds. The hydrogel system was shown to be effective against both S. aureus and E. coli pathogenic bacteria [25]. A proposed sensor carbon paste electrode modified with an O-CMC/poly (1-cyanoethanoyl-4-acryloyl thiosemicarbazide) copolymers showed an antibacterial activity against S. aureus and E. coli [26]. O-CMC is being attached to the surface of the cotton in order to improve the diffusion of the silver ions as well the loading efficacy of the nanoparticles and the whole complex is called cottonAg-II. Even after washing it for 20 times the compound still showed an antibacterial effect against both S. aureus and E. coli [27]. A double cross-linked hydrogel that had an effective antibacterial activity was made by cross-linking of O-CMC with calcium, silane, and silver nanoparticles. The prepared hydrogel system was effective in treating bacterial infections against E. coli and S. aureus and thus can be potentially used in bacteria-associated wound management [28]. A water-soluble quaternized O-CMC which was synthesized by grating the quaternary ammonium groups onto the O-CMC chains served as an antibacterial agent when silver nanoparticles were incorporated against the bacteria S. aureus [29]. A cotton fabric functionalized with the silver nanoparticle-based nanocomposite and O-CMC showed an excellent antibacterial activity against the E. coli and S. aureus. The product can be possibly used in the garments for hospitals in order to reduce the nosocomial infections [30]. In order to treat the burn wounds,

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Fig. 6 O-CMC promoting antibacterial function and aiding in wound healing

an antibacterial polysaccharide-based O-CMC hydrogel system is prepared which has the addition of eucalyptus essential oil, ginger essential oil, and cumin essential oil for repairing of the burnt skin. This system showed an antibacterial activity against S. aureus and E. coli [31]. An antibacterial hydrophilic layer consisting of OCMC grafted with silicone surface which has been pre-treated with the polydopamine. There was a decrease in the bacterial adhesion of E. coli and Proteus mirabilis [32]. O-CMC grafted with poly(vinyl pyrrolidone-iodine) which formed a complex hydrogel and microspheres revealed an accelerated rate of wound closure and potentially inhibited S. aureus bacterial infection [33] as depicted in Fig. 6 These reported studies indicated that the CMC derivatives and its composites had shown enhanced antibacterial activity.

3 Antifungal Activity of CMC The fungal cell wall consists of chitin next to the cell membrane, β-D-glucans just outside the chitin fibers, the mannoproteins as the outer layer of the fungal cell wall. The lipase alters the plasma membrane causing increase in the permeability. The Ca2+ mediates the permeability through the calcium regulator membrane fusion protein. The monosaccharide transporter assimilates or detoxifies the monomers of N-acetyl glucosamine. The dioxygenases and glutathione transferase are responsible for the reactive oxygen species and ATP production. The protein synthesis is also modified due to the exposure to O-CMC as depicted in Fig. 7. Cell membrane damage is the efficient way of preventing the resistance of drug in the fungal diseases. A negative model of the cell membrane structure formed by the

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Fig. 7 Molecular mechanism of action of O-CMC against the fungal growth

C-coordinated O- CMC-Cu complex was developed. FITC labeled complex was formed by the nucleophilic substitution reaction which can pass through the cell membrane. The treatment using the developed complex led to the alterations in the morphology of the cell membrane of Phytophthora capsici Leonian and the giant unilamellar vesicles. The unsaturated fatty acids and the important enzymes regulating the fatty acid synthesis were downregulated along with most of the membrane proteins which are needed for the substance transport and various biochemical reactions were also downregulated by the developed complex. Also, this complex caused the leakage of intercellular electrolytes and the proteins along with the sugar [34]. O-CMC was modified by graft copolymerization using various monomers to improve its properties and applications. Styrene was graft polymerized onto O-CMC using ammonium persulfate as an initiator and had antifungal activity against Candida albicans, Aspergillus fumigatus, Geotrichum candidum, and Syncephalastrum racemosum [35]. A gauze coated with O-CMC was proven to be capable enough in preventing the fungal growth of C. albicans [36]. A peptide antimicrobial in nature named Octominin was encapsulated into chitosan nanoparticles and O-CMC showed inhibition of various fungi like C. albicans and A. baumannii by cell membrane permeability and also reactive oxygen species formation and eradication of biofilm [37]. O-CMC Schiff base was chemically modified and their metal complexes like Co (II), Ni (II), and Zn (II) in order to inhibit the fungal growth against C. albicans to the maximum extent followed by A. niger and A. clavatus [38]. O-CMC Schiff bases synthesized by the condensation reaction and then coordination reaction of Cu (II) complexes cupric ions had the potency to have antifungal properties against Phytophthora capsica, Glomerella cingulate, and Gibberella zeae and were efficient than the commercial chitosan

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[39]. Another type of O-CMC Schiff bases by condensation reaction and later coordinating with cupric, nickel, and zinc ions and the metal complexes had showed the antifungal activity against Phytophthora capsica, Fusarium oxysporum, Botrytis cinerea, and Gibberella zeae [40]. O-CMC with different molecular sizes that was obtained by etherification with the use of chloroacetic acid later alkylated with the C4-C12 fatty aldehyde through the Schiff-base reaction. The resultant was derivate of the amphiphilic N-alkyl-O-CMC that has a strong inhibition against Alternaria macrospora [41]. A hydrogel was prepared using CMC and cross-linked with different concentrations of N, N′-bis (4-(isothiocyanate carbonyl) phenyl) pyremellitimide to produce pyromellitimide benzoyl thiourea-O-CMC hydrogel that is potent to inhibit various fungi. The main fungi inhibited were Aspergillus fumigatus, Geotrichum candidum, and Syncephalastrum racemosum. The hydrogel due to its hydrophilic nature because of the presence of pyromellitimide thiourea moieties enhanced the active ingredients diffusion into the pathogens causing the disturbances of the enzymes [42]. O-CMC with two different molecular weights was attached with 1-(4-(2-aminoethyl) phenoxy) zinc phthalocyanine and later quaternized which yields eight new conjugates which had inhibition of growth against Candida albicans [43]. In another study the antifungal effect of low and high molecular weight O-CMC, chitosan oligosaccharide, and N- acetyl-D- glucosamine showed activity against Candida albicans, Candida glabrata, and Candida krusei [44]. These reported studies demonstrated that CMC derivatives and its composites had shown significant antifungal activity.

4 Conclusion This chapter discusses the antimicrobial activity of CMC derivatives and their composites based on the reported studies. This water-soluble CMC derivative and their composites enhanced the physicochemical and biological properties. Owing to the modified properties of CMC, their composites showed a better antibacterial and antifungal properties. Although several in vitro antimicrobial studies have been studied and reported by several researchers, further in vivo studies are required to prove the complete antimicrobial efficacy of the CMC derivatives and their composites.

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Adv Polym Sci (2024) 292: 59–88 https://doi.org/10.1007/12_2023_150 © The Author(s), under exclusive license to Springer Nature Switzerland AG 2023 Published online: 14 July 2023

Synthesis, Properties, and Applications of Carboxymethyl Chitosan-Based Hydrogels Rui Yu and Suming Li

Contents 1 Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2 Synthesis of CMCS-Based Hydrogels . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2.1 CMCS-Based Physical Hydrogels . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2.2 CMCS-Based Chemical Hydrogels . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3 Properties of CMCS-Based Hydrogels . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.1 Morphology . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.2 Self-healing Behaviors . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.3 Antibacterial Activities . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.4 Mechanical Properties . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.5 Swelling Performance . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.6 Stimuli Sensitivity . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4 Applications of CMCS-Based Hydrogels . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4.1 Drug Delivery . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4.2 Tissue Engineering . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4.3 Wound Dressing . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5 Conclusion and Perspectives . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

60 61 61 65 69 69 71 72 73 74 75 76 76 78 79 80 82

Abstract Carboxymethyl chitosan (CMCS), as one of the most investigated derivatives of chitosan, has been widely used for the synthesis of hydrogels because of its

R. Yu Guangzhou Municipal and Guangdong Provincial Key Laboratory of Molecular Target & Clinical Pharmacology, the NMPA and State Key Laboratory of Respiratory Disease, School of Pharmaceutical Sciences and the Fifth Affiliated Hospital, Guangzhou Medical University, Guangzhou, China S. Li (✉) Institut Européen des Membranes, Université de Montpellier, Montpellier, France e-mail: [email protected]

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good water solubility, biocompatibility, and biodegradability. CMCS-based hydrogels can be obtained through physical approaches such as hydrogen bonding, freeze-thawing, and ionotropic gelation, or via chemical approaches such as grafting, free-radical polymerization, enzymatic reaction, and condensation reaction. The hydrogels exhibit outstanding physicochemical and biological properties, including interconnected porous morphology, self-healing and swelling performance, antibacterial activity, and stimuli sensitivity, which makes them a material of choice for various biomedical applications, particularly in drug delivery, tissue engineering, and wound dressing. This chapter aims to provide an overview of the latest advances in the synthesis, properties, and biomedical applications of CMCS-based hydrogels. Keywords Carboxymethyl chitosan · Drug release · Hydrogel · Tissue engineering · Wound healing

1 Introduction Hydrogels are a three-dimensional (3D) network of hydrophilic polymers with the ability to absorb large amounts of water or biological fluids without dissolving or losing their structural integrity [1]. This 3D network is formed by crosslinking polymer chains via covalent bonds (chemical hydrogel) or noncovalent bonds such as hydrogen, ionic, or hydrophobic bonds (physical hydrogel). The presence of hydrophilic groups, such as hydroxyl, carboxyl, amine, amide, or sulfonic groups, endows hydrogels with excellent water absorption capacity [2–4]. Some “smart” hydrogels are able to respond to external stimuli like pH, temperature, ions, and light [5, 6]. The porous and interconnected structure of hydrogels allows the diffusion of bioactive molecules in the network [1]. In general, hydrogels present physical characteristics similar to native extracellular matrix (ECM) or living tissue, which makes them a material of choice for biomedical applications in drug delivery, wound dressing, and tissue engineering [2–4]. Hydrogels can be obtained from synthetic polymers, such as poly(vinyl alcohol) (PVA), poly(2-hydroxyethyl methacrylate) (PHEMA), and poly(ethylene glycol) diacrylate (PEGDA), and biopolymers, such as polysaccharides (alginate, chitosan, cellulose, hyaluronic acid) and proteins (gelatin, fibrin, collagen) [7–9]. PHEMA-, PVA-, and PEGDA-based hydrogels present interesting mechanical properties, but they are not biodegradable under in vivo conditions. In contrast, biopolymer-based hydrogels exhibit outstanding biocompatibility and biodegradability, and are most promising for applications in the biomedical and pharmaceutical fields [10]. Among the various biopolymers, chitosan has received much attention for the conception of biomaterials because of its outstanding properties such as biodegradability, biocompatibility, and antibacterial properties [11, 12]. However, chitosan is insoluble in aqueous media at neutral or alkaline pH because of strong inter- and intramolecular hydrogen bonding, which greatly limits its potential applications.

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Chitosan possesses functional amino groups, as well as primary and secondary hydroxyl groups along the polymer chains, which allows chemical modification under mild reaction conditions. Many water-soluble derivatives have been prepared by introducing hydrophilic groups, such as carboxymethyl, di-hydroxyethyl, sulfate, phosphate, and (hydroxy)alkylamino, or by grafting a water-soluble polymer on the macromolecular chain of chitosan [13]. Among them, carboxymethyl chitosan (CMCS) has attracted wide interest because of its low cost, ease of synthesis, and high water solubility [3, 14]. Generally, carboxymethylation reaction occurs either on hydroxyl groups (C-6) or on NH2 groups of the D-glucosamine moieties of CMCS. The obtained derivatives contain primary amines (-NH2) or secondary amines (-NH-CH2COOH) [15]. Therefore, CMCS is usually categorized as N,O-carboxymethyl chitosan (NO-CMCS), O-carboxymethyl chitosan (O-CMCS), and N-carboxymethyl chitosan (N-CMCS). The physicochemical and biological properties of CMCS mainly originate from the miscellaneous functional groups (hydroxyl, amino, and carboxyl) on its backbone. Similar to the parent chitosan, CMCS presents excellent antibacterial ability which mainly depends on the content of positively charged -NH3+ functional groups [16]. It also possesses outstanding chelating, sorption, moisture retention, cell functioning, antioxidant, and antiapoptotic properties [17]. To date, there is a booming number of publications on CMCS-based materials and their applications in many domains, especially in biomedical and pharmaceutical domains [18–23]. This contribution aims to provide a comprehensive review of the synthesis, properties, and applications of CMCS-based hydrogels.

2 Synthesis of CMCS-Based Hydrogels The physicochemical properties of hydrogels, including the morphology, selfhealing and swelling behavior, mechanical properties, and malleability, are strongly related to the preparation approaches. Therefore, it is of great importance to select an appropriate approach to prepare CMCS-based hydrogels with desired properties. For example, self-healing hydrogels can be synthesized by introducing reversible dynamic bonds [24]. Many crosslinking methods have been developed to prepare different types of hydrogels [25–27]. Hydrogels are generally classified into two categories based on the preparation method, i.e., chemical hydrogels and physical hydrogels. In Table 1 are summarized the various CMCS-based hydrogels reported in the literature.

2.1

CMCS-Based Physical Hydrogels

Physical hydrogels have gained great interest for various applications in the fields of food, pharmaceutics, and biomedicine due to the relatively facile preparation

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Table 1 CMCS-based hydrogels obtained by physical or chemical crosslinking Composition Sodium alginate/CMCS PVA/CMCS/AgNPs/borax CMCS/oligomeric procyanidins

Approaches Hydrogen bond Hydrogen bond Hydrogen bond

Bletilla striata polysaccharide/CMCS/ Carbomer 940 CMCS/tannic acid PVA/CMCS/tannic acid PVA/human-like collagen/CMCS PVA/CMCS-N,N′-di-(L-alanine)-3,4,9,10perylene tetracarboxylic diimide/tannic acid CMCS/PVA Methoxy poly(ethylene glycol)/CMCS/ alginate Protocatechuic acid/CMCS/oxidized sodium alginate (CMCS-g-PANI)/Ag+ CMCS/NIPAAm

CMCS/acrylic acid Protocatechuic acid/CMCS Quaternized CMCS/PEG CMCS/acrylic acid CMCS/2-hydroxyethyl acrylate

Sodium 2-acrylamido-2-methylpropane sulfonate/CMCS β-cyclodextrin/CMCS/2-Acrylamido-2methylpropane sulfonic acid CMCH/catechin/laccase Silk fibroin/CMCS Oxidized hydroxyethyl cellulose/CMCS CMCS/oxidized dextran/ poly-γ-glutamic acid Oxidized quaternized guar gum/CMCS

Reference [28] [29] [30]

Hydrogen bond

Application Protein delivery Wound dressing Antibacterial material Wound dressing

Hydrogen bond Freeze-thawing Freeze-thawing Freeze-thawing

Wound dressing Wound dressing Wound dressing Wound dressing

[32] [33] [34] [35]

Freeze-thawing Grafting, condensation Grafting, condensation

Skin scaffold Drug delivery

[36] [37]

Drug release and tissue engineering Wearable devices Drug delivery

[38]

Drug delivery Tissue engineering Wound dressing

[41] [42]

Drug release Drug release

[44] [45]

Wound dressing

[46]

[31]

Grafting, electronic static Grafting, freeradical polymerization Grafting Grafting, condensation Grafting, condensation Grafting Grafting, freeradical polymerization Free-radical polymerization Free-radical polymerization Enzymatic reaction Enzymatic reaction Condensation Condensation

[39]

Drug delivery

[47]

Tissue engineering Cartilage scaffold Wound dressing Wound dressing

[48]

Condensation

Wound dressing

[40]

[43]

[49] [50] [51] [52] (continued)

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Table 1 (continued) Composition Oxidized konjac glucomannan/CMCS/ graphene oxide Gold nanoclusters/CMCS/oxidized carboxymethyl cellulose CMCS/PNIPAm/glycidyl methacrylate

Approaches Condensation

Application Wound dressing

Reference [53]

Condensation

Biosensor

[54]

Condensation

Drug delivery

[55]

Fig. 1 Preparation of sodium alginate/CMCS hydrogel beads by dropping a solution of the two polymers into a citric acid solution [28]

approach without using any crosslinkers. Physical crosslinking is achieved by physical interactions between polymer chains, including hydrogen, ionic, or hydrophobic bonds.

2.1.1

Hydrogen Bonding

Hydrogen bonding is a key feature of chemical structure and reactivity [56]. It refers to inter- or intramolecular interactions which involve a hydrogen atom adjacent to two other highly electrophile atoms. The hydrogen bond is responsible for many of the physical and chemical properties of polymers such as solubility and mechanical properties. In particular, it can serve as crosslinking domains to prepare noncovalent crosslinked hydrogels. Jing et al. reported the construction of sodium alginate/CMCS hydrogel beads via hydrogen bonding and electrostatic interactions (Fig. 1) [28]. The as-prepared hydrogel beads exhibit pH-sensitive swelling behavior and excellent lactoferrin loading capacity. Lactoferrin structure remained intact during drug loading and

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drug release processes, which suggests that the hydrogel beads could be utilized as a protein drug carrier for oral delivery. Liu et al. prepared a hydrogel composed of borax crosslinked PVA and CMCS [29]. Hydrogen bonds and borate ester bonds between polymers endow the hydrogel with outstanding self-healing behavior and mechanical properties. The as-prepared hydrogel exhibits excellent antibacterial ability due to the natural antimicrobial property of CMCS. Moreover, introduction of highly stable silver nanoparticles (AgNPs) to the hydrogel can effectively inhibit the growth of Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus), showing its potential for applications in wound healing.

2.1.2

Freeze-Thawing

Freeze-thawing is a commonly used method for preparing CMCS hydrogels due to its simplicity and efficiency. The freeze-thawing process involves freezing the solution followed by thawing at room temperature. During freezing, ice crystals develop and expand, thus concentrating the solutes in the remaining liquid phase. Upon thawing, the solutes form a network structure as a result of the increased concentration, which leads to the formation of a hydrogel network [57]. Zhou et al. constructed a composite hydrogel from PVA, CMCS, and tannic acid (TA) by using cryogenic treatment and freeze-drying method [33]. The synthesis approach was facile and feasible for formulating hydrogel dressings. Pourjavadi et al. synthesized a double network hydrogel based on O-CMCS, PVA, and graphene oxide [36]. CMCS chains crosslinked by CaCl2 serve as the first network, whereas PVA crosslinked by freeze-thawing serves as the second network. CMCS exhibits good miscibility with PVA in the presence of honey used as plasticizer. The addition of graphene oxide reinforces the polymeric network, and thus the as-prepared hydrogels exhibit good tensile strength. Besides, the hydrogels present good cell viability and biodegradability, and can be used as skin scaffold. The characteristics of CMCS-based hydrogels prepared by freeze-thawing can be adjusted by varying the freezing and thawing parameters, such as freezing speed, thawing speed, and the number of freeze-thawing cycles, which is promising for a wide range of applications.

2.1.3

Ionotropic Gelation

Ionotropic gelation is one of the most widely used methods to prepare chitosan- or CMCS-based hydrogels [58]. Typically, CMCS solution is mixed with an ion crosslinking agent, such as sodium tripolyphosphate or calcium chloride, to form a hydrogel network through electrostatic interactions. The ion crosslinking agent reacts with the carboxyl groups of CMCS, resulting in ionic bonding and hydrogel formation. Chu et al. synthesized a CMCS-based hydrogel from CMCS and different zinc salts, including ZnSO4, Zn(NO3)2, and ZnAc2. Data showed that CMCS hydrogel

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exhibits higher fluorescence in the presence of Zn2+ ions due to complexation of Zn2+ with carboxyl, amine, and hydroxyl groups [59]. The authors suggested that these hydrogels could find applications in wound dressing, drug delivery, and bioimaging [60]. Huang et al. synthesized composite hydrogel beads based on polyacrylamide-modified kaolin, sodium alginate, and CMCS for adsorption of heavy metals such as Cu2+ [61]. The results showed that the adsorption on hydrogel beads conforms to the quasi-second-order kinetics and Langmuir isotherm models, providing a reference for improving the adsorption performance of biomass-derived adsorbents in the field of sewage treatment. The properties of CMCS-based ionotropic hydrogels can be adjusted by changing the concentration of CMCS and ions, as well as the crosslinking time and temperature. Overall, the ionotropic gelation method is a simple and efficient technique for producing CMCS-based hydrogels with desired properties. Moreover, its scalability and reproducibility make it an attractive choice for large-scale production.

2.2

CMCS-Based Chemical Hydrogels

Physical hydrogels are generally brittle and unstable. Chemical crosslinking methods are thus used to construct hydrogels with better stability and mechanical properties. A variety of chemical crosslinking approaches have been developed, including grafting, free-radical polymerization, condensation, enzymatic reaction, and addition reaction.

2.2.1

Grafting

Chemical hydrogels can be obtained by attaching side chains onto the backbone of hydrophilic polymers such as polysaccharides, including chitosan, cellulose, alginate, and their derivatives. The polymeric backbone can be activated by the action of high energy radiation or chemical reagents. The polymerization of functional monomers on activated backbones results in branching and finally to the formation of a network. Zhang et al. synthesized a dual-network conductive hydrogel by combining covalently bonded polyacrylamide (PAM) and ionically crosslinked Ag+/CMCS-gpolyaniline (Ag+/CMCS-g-PANI) [39]. The amino groups on CMCS chains serve as the active sites to initiate polymerization of aniline for the grafting of PANI. The resulted hydrogel presents remarkable biological, mechanical, and electronic properties such as biocompatibility, antibacterial activity, elasticity, adhesiveness, and sensitivity for strain sensors, and could find applications in wearable electronic devices. Khan et al. reported the preparation of in situ crosslinkable hydrogels based on poly(N-isopropylacrylamide) (PNIPAAm) and CMCS [40]. The developed hydrogels exhibit dual pH/thermo-sensitivity, which is promising for uses in systemic and intratumoral controlled drug delivery.

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Free-Radical Polymerization

Free-radical polymerization is also a commonly used method to prepare CMCSbased hydrogels. CMCS can react with polymers modified with functional groups which are prone to free-radical polymerization by the action of an initiator such as azobisisobutyronitrile (AIBN) and 4,4′-azobis(4-cyanovaleric acid) (ACVA), and a crosslinking agent such as N,N′-methylenebisacrylamide (MBA). Rodkate et al. synthesized multi-responsive composite microspheres via in situ free-radical polymerization of NIPAAm in the presence of CMCS and magnetite nanoparticles (MNPs) followed by glutaraldehyde crosslinking (Fig. 2) [62]. Thermal- and pH-responsive drug release was obtained from the microspheres, indicating their potential for use in controlled release applications. Jeong et al. prepared a pH-sensitive hydrogel via radical polymerization of 2-hydroxyethyl acrylate (2-HEA) from CMCS chains, using MBA as crosslinking agent [45]. Typically, the initiator (potassium persulfate, KPS) produces amino radicals along CMCS chains, and the radicals initiate the polymerization of 2-HEA and MBA. The as-prepared hydrogels exhibit outstanding viscoelasticity and porous structure, and can be applied as a pH-sensitive transdermal drug delivery system due to the pH sensitivity of CMCS. The properties of hydrogels can be adjusted by changing the CMCS concentration, the crosslinking agent, the initiator, and the polymerization time and temperature.

2.2.3

Enzymatic Reaction

Enzymatic reaction is an effective method to prepare CMCS-based hydrogels because it can be performed under mild conditions and is highly selective. Andreia et al. synthesized CMCS-based hydrogels via enzymatic crosslinking with catechin, one of the most commonly studied polyphenols, in the presence of a polyphenol oxidase, i.e., laccase [48]. Rheological and structural measurements revealed the formation of crosslinks in the resulting hydrogel. Li et al. developed silk fibroin (SF) and CMCS composite hydrogels with enzyme-mediated chemical crosslinking (horseradish peroxidase and hydrogen peroxide) and β-sheet crosslinking (ethanol

Fig. 2 Synthesis of composite hydrogel microspheres via in situ free-radical polymerization of NIPAAm in the presence of CMCS and magnetite nanoparticles (MNPs) followed by glutaraldehyde crosslinking [62]

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Fig. 3 Synthesis of silk fibroin (SF) and CMCS composite hydrogel in two steps: (a) attachment of tyramine (TA) to CMCS using EDA/NHS as coupling agents, (b) formation of SF/CMCS hydrogel via enzyme-mediated chemical crosslinking with horseradish peroxidase (HRP) and hydrogen peroxide (H2O2) and β-sheet crosslinking with ethanol treatment [49]

treatment) (Fig. 3) [49]. Both CMCS-based hydrogels present good biocompatibility and tailorable physicochemical properties and could be used as scaffolds in regenerative medicine. A dual-network hydrogel was synthesized from CMCS and oxidized methacrylate alginate. CMCS was first functionalized with collagen peptide in the presence of glutamine transferase [63]. Hydrogel was then obtained by Schiff base reaction between CMCS and oxidized methacrylate sodium alginate, followed by photocrosslinking of methacrylate groups under UV light. The as-prepared hydrogel with double network exhibits interconnected porous morphology, good mechanical properties, and outstanding biocompatibility. A mouse full-thickness skin wound model was used to evaluate the potential of the hydrogel in promoting wound healing. Data showed that the hydrogel could regulate the inflammatory process, enhance collagen deposition, and improve vascularization, demonstrating its potential for wound dressing application.

2.2.4

Condensation Reaction

Polysaccharides have many carboxyl, hydroxyl, or amine groups along the backbone. These functional groups can be used to synthesize chemical hydrogels via condensation reaction. For example, hydrogels can be obtained by controllable crosslinking of multivalent polymers via Ugi four-component reaction [64]. Condensation between amines and aldehydes or ketones leads to the formation of an imine bond plus a water molecule. This is a classical reaction in organic chemistry discovered in 1864 by Hugo Schiff, a German chemist [65]. A Schiff base has a chemical structure of R1R2C=NR3, where R1 and R2 represent an aryl group, an alkyl group, or a hydrogen, and R3 an alkyl or aryl group [66]. Schiff base reaction is reversible. Many factors, such as concentration, temperature, pH, electronic, or steric

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Fig. 4 Equilibrium-controlled reactions of imine bonds: (a) condensation/hydrolysis, (b) exchange of the two R groups by addition of an amine, and (c) metathesis—exchange of the two R groups upon introduction of a second imine

Fig. 5 Synthesis route of CMCS-based dynamic hydrogel by Schiff base reaction in two steps: (1) reaction between equimolar Jeffamine and BTA to yield a dynamer (Dy) and (2) reaction between Dy and CMCS to yield a hydrogel network

effect, can influence the equilibrium of the reaction. In principle, an imine bond can undergo three types of equilibrium-controlled reaction [66], i.e., hydrolysis, exchange, and metathesis. In Fig. 4 are illustrated these three reactions involving imines: (a) hydrolysis of the imine bond could occur in the presence of water, (b) the two R groups can be exchanged by addition of an amine, and (c) the two R groups can be exchanged upon introduction of a second imine. In a comprehensive study on CMCS-based dynamic hydrogels, Yu et al. synthesized a series of hydrogels from CMCS and Jeffamine by Schiff base reaction in two steps, as shown in Fig. 5. First, Jeffamine with an amino group at both ends reacts with a crosslinker, i.e., benzene-1,3,5-tricarbaldehyde (BTA), yielding a watersoluble dynamer (Dy). Second, reaction between Dy and CMCS yields a hydrogel network. The effects of the composition and the molar masses of Jeffamine and CMCS on the hydrogel properties were investigated. The hydrogels present outstanding self-healing performance, pH-sensitive swelling, excellent cytocompatibility versus mesenchymal stromal cells (MSCs), and remarkable

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antibacterial activity against E. coli, a gram-negative bacterium [67, 68]. Thymopentin (TP5), a hydrophilic immunostimulant, was added in one of the precursor solutions before gelation. Drug release studies showed a burst release followed by a slower and prolonged release, and the release rate is mainly dependent on the drug loading and release media. These findings demonstrate the great potential of dynamic hydrogels in regenerative medicine, wound healing, and drug release [69]. Dynamic hydrogels can also be obtained via Schiff base reaction between oxidized alginate (oALG) or oxidized hyaluronic acid (oHA) with CMCS. In fact, aldehyde groups can be introduced to ALG and HA via oxidation of uronic acid units by sodium periodate (NaIO4). The two hydroxyl groups on C2 and C3 positions of β (1 → 4) linked uronic acids are oxidized to aldehydes, thus yielding dialdehyde derivatives oALG or oHA. It is noteworthy that oxidation can occur for both mannuronic acid and guluronic acid units of ALG, while only the glucuronic acid units can be oxidized in the case of HA. Hydrogels can be prepared by crosslinking aldehyde groups of oALG or oHA with amine groups present in CMCS [70, 71]. Nguyen et al. prepared hydrogels from CMCS and oHA with different oxidation degrees [72]. Data showed that the CMCS/oHA volume ratio directly affects the homogeneity, porosity, and degradation behavior of the hydrogels. Lower oxidation degree of oHA favors cell proliferation, cell attachment, and wound healing process [72]. Mohabatpour et al. prepared hydrogels at varying weight ratios of oALG to CMCS through Schiff base reaction [73]. The resulted hydrogels exhibit rapid self-healing ability after injection, good antibacterial activity against cariogenic bacteria, and high viability of HAT-7 cells, thus showing great potential in cell delivery and injection cell therapy.

3 Properties of CMCS-Based Hydrogels 3.1

Morphology

Hydrogels are three-dimensional networks of hydrophilic polymers that possess unique properties, making them promising materials for various applications. One of the key parameters that affects the properties of hydrogels is their morphology, which can be precisely controlled by various factors, including the composition, the synthesis method, and the crosslinking density. Yu et al. synthesized hydrogels by crosslinking CMCS and diamino Jeffamine in the presence of BTA via Schiff base reaction [69]. Three hydrogels (Gel80K, Gel30K, and Gel25K) were obtained from CMCS with molar mass of 80,000, 30,000, and 25,000, respectively. A highly porous and interconnected structure was observed in all cases (Fig. 6). The average pore diameter of Gel80K, Gel30K, and Gel25K is 82.9 μm (Fig. 6a, b), 76.1 μm (Fig. 6c, d), and 59.5 μm (Fig. 6e, f), respectively, showing the effect of the molar mass of CMCS on the pore size of the hydrogel network. On the other hand, the crosslinking density plays a major role in

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Fig. 6 Pore structures and pore size distributions of CMCS-based dynamic hydrogels: Gel80K (a, b), Gel30K (c, d), and Gel25K (e, f) [69]

tuning the pore size of hydrogel network. Lin et al. synthesized hydrogels by imine bonding between CMCS and oxidized hydroxyethyl cellulose (oHEC) at different oxidation degrees [50]. Data showed that with the increase of the degree of oxidation, the 3D network structure of hydrogels becomes denser and the number of pores decreases due to higher crosslinking density. Chen et al. conceived a hydrogel from CMCS, oxidized dextran, and poly-γ-glutamic acid (γ-PGA) with the formation of

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dynamic imine bonds, intermolecular amide bonds, and intramolecular lactam bonds [51]. The authors observed that the pore size and distribution are dependent on the crosslinking density of hydrogels. Higher crosslinking density leads to smaller pore size. Yu et al. synthesized a hydrogel from CMCS and oxidized quaternized guar gum (oQGG) [52]. The authors also found that with increased amount of aldehyde groups in oQGG, the degree of crosslinking of hydrogels became higher and the hydrogel structure became denser.

3.2

Self-healing Behaviors

Another remarkable characteristic of CMCS-based hydrogels is their ability to selfheal, which allows them to effectively restore damage resulting from external forces. This is particularly true for CMCS-based dynamic hydrogels reversibly crosslinked by imine bonding [67]. In a self-healing test (Fig. 7), transparent and yellow hydrogel samples were cut into two equal parts. Then one transparent part and one yellow part were placed together at 37°C. After 20 min contact, both parts became a merged piece which could be peeled off and suspended. Geng et al. synthesized injectable hydrogels via mixing solutions of TA and CMCS using 1,4-benzenediboronic acid (BDBA) as crosslinker [74]. Gelation occurs via hydrogen bonds between TA and CMCS, as well as via dynamic boronate ester bonds between TA and BDBA. The as-prepared hydrogel is able to self-heal via reconstruction of reversible bonds [74]. Zhao et al. constructed an electroactive hydrogel from oxidized sodium alginate, CMCS, and silver nanoparticles [75]. The hydrogel exhibits a good injectability, tunable gelation time, and good self-healing performance. These studies demonstrated the exceptional self-healing capacity of CMCSbased hydrogels, opening up a new and promising avenue of research for biomaterials with improved mechanical properties and extended lifespan.

Fig. 7 Self-healing test of CMCS-based dynamic hydrogels prepared from CMCS, diamino Jeffamine, and benzene-1,3,5-tricarbaldehyde through Schiff base reaction [67]

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Antibacterial Activities

Chitosan possesses inherent antibacterial activity due to numerous amino groups present along polymeric chains. Liu et al. reported that the parent chitosan and CMCS derivatives present an increasing order of antibacterial activity against E. coli: NO-CMCS70% of the drug was delivered in 20 h. The enhanced solubility of the CaCO3 in the hybrid NPs at a reduced pH was the main reason of the pH-dependent releasing behavior. Due to the acidic tumor microenvironment, this drug release profile is advantageous to induce faster drug delivery at tumor site. Maya et al. [11] developed O-CMC-Cetuximab conjugated NPs for tumortargeted drug delivery of paclitaxel. These NPs showed a controlled and pH-dependent paclitaxel release profile due to their pH-responsive swelling behavior. At alkaline medium, the developed conjugates exhibited the slow rate of drug release due to surface conjugation of Cetuximab with the polymer matrix. The results of the hemolysis assay revealed a reasonable hemolytic ratio and unbroken RBCs, suggesting the non-hemolytic activity of NPs. Cancer cell lines such as A549, A431, and SKBR3 treated with the developed NPs showed 80% cell death. Fluorescent microscopy and flow cytometry analyses revealed that the prepared NPs showed an increased cellular uptake by EGFR overexpressed cancer cell types. Small interfering RNAs (siRNA) have been loaded into self-assembled CMC conjugated with fluorescein isothiocyanate (FITC)-labeled chitosan hydrochloride NPs as pH-responsive or mucosa-targeted epithelial drug delivery systems [12]. The encapsulation effectiveness of siRNA in the NPs was 76.7% at the ideal condition. These NPs presented the regulated release of siRNA by responding to an external stimulus under favorable pH values while protecting siRNA from the deprivation by acidic media. These NPs were found to be suitable for delivering siRNA into colon cancer cells.

2.1.2

Temperature-Dependent Drug Delivery

A range of temperature and pH-responsive nanogels based on poly (N-isopropylacrylamide-co-2-acrylamido-2-methyl-1-propanesulfonate-co-1propene-2-3-dicarboxylate) conjugated with N,O-CMC was prepared for tumortargeted drug delivery [13]. At 37°C and pH 5.4, these nanogels exhibited a swelling equilibrium and phase transition behavior to govern the controlled drug release profile. These drug-loaded nanogels showed an increased cytotoxicity against mammary cancer cells compared to normal epithelial cells due to their tumortargetability. Moreover, these nanogels effectively aggregated in MCF-7 cells and caused DNA damage and mitochondrial-facilitated death. For the treatment of cancer, the use of these drug carriers can be expanded to a wide spectrum of positively charged pharmaceuticals. They are also a prime choice for in vivo drug delivery due to their thermos-responsiveness.

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2.1.3

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Charge Inversion Response Conjugate

In a recent study, DOX was encapsulated in the cavity of a polyamidoamine (PAMAM) dendrimer and then PAMAM was modified by CMC through electrostatic interaction to produce a nanocarrier that exhibits charge inversion response to tumor microenvironment for targeted anticancer drug delivery (Fig. 4) [14]. Under physiological settings, the nanocarrier was negatively charged with the protection of CMC; however, when the covering material was removed to treat an acidic tumor, the complex became positively charged, facilitating cellular uptake through electrostatic absorptive endocytosis. The developed nanocarrier showed a 1.5-fold better anticancer activity after intravenous administration in mice with H22 tumors than free medicines did, and histological investigation revealed no clear systemic harm.

Fig. 4 Schematic representation of CMC-modified PAMAM dendrimer

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CMC-Drug Conjugates

Chi et al. created novel CMC-norcantharidin conjugates for cancer therapy [15]. These conjugates caused SGC-7901 cells to undergo apoptosis and dramatically slow down cell proliferation. In BALB/c nude tumor-bearing mice, the developed CMC-norcantharidin conjugates had better anticancer activity than free drug with a tumor inhibition rate of 59.57%. The enhanced antitumor effects may be caused by increased expression of TNF-α (mouse tumor necrosis factor-α) and decreased expression of CD34, VEGF, MMP-2, and MMP-9. Additionally, it was shown by western blot assay and immunohistochemical analysis that the conjugates caused cell death by suppressing the Bcl-2 expression and enhancing the expressions of Caspase-3 and Bax.

2.1.5

CMC-Modified with Graphene Oxide (GO)

Yang et al. [16] developed DOX-loaded GO modified with CMC followed by conjugation of hyaluronic acid (HA) and FITC. The developed complex specifically targeted cancer cells that overexpress with CD44 receptors and effectively blocked their growth. At acidic pH, the drug loading capacity and release rate of the complex were found to be higher than that at pH 7.4. The complex showed an increased cytotoxicity against Hela cells compared to L929 cells. This shows that the receptormediated binding and intracellular uptake of the complex enable targeted suppression of the development of cancer cells.

2.1.6

CMC-Iron Oxide Nanocarrier

Li et al. [17] produced core–shell nanocarriers that consist of iron oxide (Fe3O4) @SiO2 as a core and CMC conjugated with folic acid (FA) as a shell for tumor targeted drug delivery (Fig. 5). Due to the presence of FA, the developed nanocarriers were found to be absorbed by HeLa cells within a short period of time. It was observed that after cellular uptake, the nanocarriers were primarily distributed in the cytoplasm. These benefits make the produced core–shell nanocarriers attractive for cancer-specific targeting and therapy.

Fig. 5 Synthesis of Fe3O4@SiO2-CMC-FA nanocarrier

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CMC Coated with Gold (Au) NPs

Kang et al. [18] synthesized more stable DOX-encapsulated Au-coated CMC for cancer therapy. In this work, CMC was used as a capping and stabilizing agent. In another study, DOX was immobilized on Au NPs and covered with CMC for efficient delivery to cancer cells [19]. These particles presented the tumor-targeted delivery of DOX at acidic medium due to the ionization of carboxylic group. In comparison to free DOX, the DOX-loaded Au NPs were effectively taken up by cancer cells and their uptake increased under acidic condition. In addition, these NPs were found to be stable over a wide pH and electrolyte concentration range.

2.2

Organ-Specific Drug Delivery

Compared to other drug delivery methods, organ-specific drug delivery has a number of advantages, including less side effects, increased therapeutic effects, bioadhesiveness, prolonged release potential, and ease of administration [20]. Numerous researchers have developed CMC-based formulations that solely carry certain drugs to specific organs such as eyes, gut, liver, pancreas, colon, and others for their activity.

2.2.1

Intestine-Targeted Drug Delivery

Huang et al. [21] prepared glutaraldehyde-crosslinked O-CMC/gum arabic (GA) for the delivery of drugs at intestine target. The carriers showed diverse swelling ratios in the intestinal media and released majority of the drug into the simulated gut and colon solutions. In another study, bovine serum albumin (BSA)-encapsulated OCMC-GA microcapsules were created at acidic media (pH 3.0, 4.5, and 6.0) using genipin as a crosslinking agent. The stability of these microcapsules was found to be more at pH 6.0 than that at pH 4.5. Hence, these microcapsules could be suitable for transferring pharmaceuticals to the colon site [22].

2.2.2

Colon-Targeted Drug Delivery

The third largest cause of mortality worldwide is colorectal cancer. Chemotherapy is still the only effective method for treating colorectal cancer. Improved anticancer efficacy and targeted distribution of therapeutic agents to the colon remain difficult for the colorectal cancer treatment. It was found that mucoadhesive nanocarriers have increased anticancer activity with a tailored drug delivery ability. The unique surface properties of mucoadhesive nanocarriers can improve cellular absorption of formulation by increasing the interaction between drugs and intestinal mucosa.

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Increased local drug concentration is produced in the lesion, leading to increased cancer cell death, which helps to minimize the dosage and frequency of administration as well as the cost and toxicity. Recently, chitosan-CMC-based mucoadhesive nanogels were prepared for colorectal cancer therapy [23]. Two types of DOX-loaded nanogels with opposing zeta potential were produced using tripolyphosphate (TPP) and CaCl2, respectively. The nanogels crosslinked by CaCl2 were more thoroughly absorbed by colorectal cancer cells than the gels crosslinked with TPP. The mucoadhesion and permeability of CaCl2 crosslinked nanogels were found to be improved due to their high mucin binding capacity and reduced paracellular transport by colon. This helps to increase local drug concentration and extend the period that a formulation is in touch with the gut mucosa. In another study, CMC/acrylic acid hydrogel formulations with different amounts of CMC, acrylic acid, and ethylene glycol dimethacrylate (EGDMA) were made by free radical polymerization [24]. The swelling and the release of 5-Fluorouracil from the hydrogel increased with increasing acrylic acid concentration while decreasing with rising EGDMA concentration. At higher pH levels, CMC helps enhance swelling and drug release due to its carboxylic groups. It was shown that raising the concentration of acrylic acid and CMC increased the gel fraction and porosity. It was determined that the release of 5-Fluorouracil increased with an increase in acrylic acid concentration whereas the release of 5-Fluorouracil decreased with an increase in EGDMA ratio. The results showed that the developed hydrogel is suitable for the delivery of 5-Fluorouracil at the colon site. A pH-sensitive composite hyaluronic acid/gelatin hydrogel incorporated with curcumin-loaded CMC was developed for inflammatory bowel disease (IBD) [25]. It remained for a considerable amount of period in the gastric fluid and dramatically swelled in intestinal fluid. The release of curcumin from the hydrogel was minimal (about 65%) in simulated gastric medium for 50 h. In vivo studies presented that the developed hydrogel lowered the serum levels of pro-inflammatory factors and preserved colon tissue. Pharmacokinetic tests and ex vivo fluorescence imaging studies showed that the hydrogel formulation sustained a high level of curcumin in colon tissue for a considerable amount of period. Since the prepared hydrogel increased the bioavailability of curcumin, it could be suitable for treatment of IBD.

2.2.3

Ocular-Targeted Drug Delivery

The most popular form of treatment for glaucoma is topical drug delivery. The thin tear layer, which is always replenished by reflex lachrymation and lid blinking, generally lowers the ocular bioavailability to