Effect of the molecular architecture on the thermosensitive properties of chitosan-g-poly(N-vinylcaprolactam)

Highlights

• Chitosan-g-(N-vinylcaprolactam) thermosensitive copolymers were prepared.

• These copolymers were readily soluble in water at neutral pH below the LCST.

• The LCST of these copolymers depends on the molecular architecture.

Abstract

A series of thermoresponsive copolymers based on chitosan-g-poly(N-vinylcaprolactam) were synthesized by amidation reaction using 4-(4,6-dimethoxy-1,3,5-triazin-2-yl)-4-methylmorpholinium chloride as coupling reagent. The effect of molecular architecture on the thermoresponsive properties of the graft copolymers solutions was studied by varying the chain length of the grafted poly(N-vinylcaprolactam), PVCL, (in the range from 4 to 26 kDa) and the spacing between grafted chains onto the chitosan backbone. The most interesting characteristic of these copolymers is their solubility in water at temperatures below their lower critical solution temperature (LCST). These solutions presented a LCST between 36 and 44 °C, which decreases with the spacing and length of grafted PVCL chains onto the chitosan backbone, in contrast with the limited decrease of the LCST of PVCL above a critical ${{\displaystyle \overline{\text{M}}}}_{\text{n}}$ value around 18 kDa. This behavior offers tangible possibilities for the preparation and application of sensitive bioactive formulations and “smart” drug delivery systems.

Introduction

Smart polymers have the ability to undergo reversible changes, either physical or chemical, in their properties due to external stimuli as the temperature, pH, ionic strength, light, etc. Temperature-responsive polymers may exhibit different reversible mechanisms, such as shape changes and phase transitions. These type of materials have attracted high scientific interest and possess great potential of application on diverse areas, such as bioseparation, controlled drug delivery, tissue engineering, and other applications in biomedical fields (Aguilar & San Román, 2014; Hoogenboom, 2014).

Water-soluble polymer molecules consist of both polar and non-polar hydrocarbon moieties, which interact with water molecules via hydrogen bonding or Van der Waals forces respectively. Thus, temperature-responsive polymers can exhibit a lower critical solution temperature (LCST) in aqueous media as a consequence of the overall changes in the hydrophilic–hydrophobic balance (Tager, Safronov, Sharina, & Galaev, 1993). In this sense, poly(N-vinylcaprolactam) (PVCL) is one of the well-known thermoresponsive polymers, which is non-ionic, biocompatible, water-soluble and undergoes a phase separation about 37 °C. PVCL has relatively high resistance to hydrolysis and it does not produce toxic low-molecular-weight amines during hydrolysis. Moreover, cytotoxicity assays performed with PVCL samples (molecular weights above 300 kDa) reveal that they were well tolerated in the analyzed cell cultures at concentrations below 10.0 mg mL⁻¹ (Vihola, Laukkanen, Valtola, Tenhu, & Hirvonen, 2005). This polymer exhibits a “classical” Flory–Huggins thermoresponsive phase behavior in water (Type I systems) and therefore, the value of its LCST is strongly dependent on molecular weight and concentration (Beija, Marty, & Destarac, 2011; Meeussen et al., 2000). The LCST of PVCL is also affected by the molecular dispersity or the nature of end groups (Maeda, Nakamura, & Ikeda, 2002; Sun & Wu, 2011).

PVCL prepared by conventional free radical polymerization is generally polydisperse $\left({{\displaystyle \overline{M}}}_{\text{w}}/{{\displaystyle \overline{M}}}_{\text{n}}>2.0\right)$ (Liu, Debuigne, Detrembleur, & Jérôme, 2014). Recently, low polydisperse PVCL has been obtained by controlled radical polymerization using different chain transfer agents. Some techniques, such as reversible addition–fragmentation chain transfer (RAFT) (Medeiros, Barboza, Giudici, & Santos, 2013; Ponce-Vargas, Cortez-Lemus, & Licea-Claveríe, 2013), atom transfer radical polymerization (ATRP) and macromolecular design via the interchange of xanthate (MADIX) (Beija et al., 2011; Wan, Zhou, Pu, & Yang, 2008) have been applied for the preparation of PVCL of predictable molar mass with decreasing dispersity of the macromolecular chains.

Chitosan (Cs) is a linear aminopolysaccharide obtained by extensive deacetylation of chitin. It is mainly composed of two kinds of structural units: 2-amino-2-deoxy-d-glucose and N-acetyl-2-amino-2-deoxy-d-glucose linked by a β(1 → 4) bond (Scheme 1B). Its biodegradability, biocompatibility and low toxicity make it a promising material for biomedical applications. It could be chemically modified, via its amino (C2), or primary (C6) and secondary (C3) hydroxyl groups, to achieve molecular structures for different purposes (Peniche, Goycoolea, & Argüelles-Monal, 2008).

In general, the graft copolymers can be obtained by several methods of synthesis, such as “grafting from”, “grafting through” and “grafting onto”, which involve a series of the side chains covalently bonded to a linear backbone. The “grafting onto” is a versatile technique that consists in a coupling reaction between end-functional groups of the grafted chains onto pendant functional groups of the backbone chain (Zhang & Müller, 2005). In this regard, the preparation of chitosan-graft-PVCL copolymers is documented (Prabaharan, Grailer, Steeber, & Gong, 2008; Rejinold, Chennazhi, Nair, Tamura, & Jayakumar, 2011; Rejinold et al., 2015; Rejinold, Muthunarayanan, et al., 2011; Rejinold et al., 2014). These reports account for the conjugation of the amine groups of chitosan with COOH-end PVCL using the pair EDC/NHS as coupling agent without considering the molecular weight of the grafted PVCL. The obtained products showed viability for biomedical applications due to their thermal behavior, biocompatibility and no-toxicity.

Recently, comparative studies between 4-(4,6-dimethoxy-1,3,5-triazin-2-yl)-4-methylmorpholinium chloride (DMTMM) and EDC/NHS have revealed that the former one promotes favorable characteristics for coupling of the carboxylic groups and water-soluble amine-polysaccharides. These studies proved that the use of DMTMM entails some benefits such as one-step amide bonding reactions, capacity for being used in a wide range of pH, compatibility with several bioconjugation systems and ease of removal of the reaction residues (D'Este, Eglin, & Alini, 2014; Reyes-Ortega et al., 2013).

The aim of the present research was to obtain a set of chitosan-graft-PVCL copolymers with controlled molecular architecture by varying grafted PVCL chain length as well as the degree of grafting. To this end, COOH-end PVCL homopolymer samples with different molecular weight were synthesized by means of controlled radical polymerization employing well-known transfer agents (e.g., solvent, thioacetic derivative), followed by their grafting onto the chitosan backbone using DMTMM. The structure of the graft copolymers was characterized and the effect of the molecular architecture on the thermoresponsive behavior of the aqueous solutions evaluated.