Structural and mechanical behavior of type-I collagen fibrils in presence of induced electrostatic interactions through ionic liquids.

Tuning the self-assembly of collagen has broad applications in the biomedical field owing to their desired biological performance as collagenous materials with tunable functionalities can further determine cellular responses. In this work, an attempt has been made to tune the self-assembly of collagen using ionic liquids, viz., imidazolium chloride (IC) and choline dihydrogen phosphate (CDHP) at its physiological pH, followed by probing assembled systems using various characterization methods. Turbidity measurements of fibrillar networks were performed to ascertain the rate of fibril formation in addition of imidazolium chloride and choline dihydrogen phosphate to collagen at physiological pH. Morphological changes were examined using Scanning Electron Microscope (SEM), binding affinities were measured by Microscale Thermophoresis (MST), in addition to, changes in the shear viscosity, mechanical strength of collagen fibrils when interacted with imidazolium and choline based ILs were carried out using rotational rheometer and Quartz Crystal Microbalance (QCM) measurements. Experimental result depicts that CDHP imparts better crosslinking as well as mechanical strength compare to IC, which is already known for destabilizing the triple helix structure is inhibiting the fibril formation. This self-assembled, ionic-liquid treated collagen-fibrillar system would accelerate various force modulated fibrillar network study, for mimicking the ECM and tissue engineering application.

Extraction of dermatan sulfate using ionic liquid-assisted enzymatic digestion: An efficient approach.

Dermatan sulfate is one of the major glycosaminoglycan (GAG) present in the animal hides, which is a waste/byproduct from meat industry. Efficient utilization of these meat industry wastes is garnering attention because these wastes render a possibility for their conversion into useful products. With the increased concerns over health, various initiatives have been developed to permit more efficient utilization of these by-products and thereby directly impacting environmental sustainability. Herein, we demonstrate for the first time an efficient and environmentally safe ionic liquid-assisted enzymatic process for the extraction of dermatan sulfate from buffalo hides. Dermatan sulfate has been extracted, separated, and purified from the GAG mixture using IL-assisted enzymatic digestions and chromatographic separations. NMR, FT-IR, and ESI-MS measurements showed typical characteristic peaks for dermatan sulfate. The advantages of this eco-friendly process adopted include i) use of fewer chemicals, ii) elimination of harsh chemicals, iii) elimination of various steps and sub-steps, iv) reduction in process time (12 h), and v) increase in extraction yield by 75% when compared to conventional enzymatic process (57%). Thus, the use of ionic liquids alongside enzymes will serve as an efficient methodology for the futuristic development of these derived GAGs for their potential applications.

Insights into protein-ionic liquid interaction: A comprehensive overview on theoretical and experimental approaches.

Owing to highly tunable nature, ionic liquids are nesting stance in the scientific community for a wide variety of applications ranging from electrochemistry to product purification, from chemical and biomedical applications to biotechnological interventions and proteomics. Proteins are unstable in its native form and several attempts have been made to stabilize them by addition of various additives. This review focusses on the studies conducted to improve protein stability with ionic liquids along with an emphasis on the mechanism of interaction. This review also specifies and discusses about the brief introduction to ionic liquids, evolution of first-, second-, and third generation of liquids over the years and their selection criterion and applications. Though, there are several elegant reviews available on proteins-ionic liquids interaction, this review systematically highlights the effect of ionic liquids viz., imidazolium, ammonium, phosphonium and choline-based ionic liquids (amino acid-based anions & classical anions) on fibrous proteins viz., collagen and keratin and globular proteins viz., bovine serum albumin and cytochrome c. Thus, this review elaborates the thorough investigations conducted to explore the stabilizing properties of ionic liquids over fibrous and globular proteins.

Stability of collagen in ionic liquids: Ion specific Hofmeister series effect.

In protein-ionic liquids (ILs) interactions, anions play an important role. In this work, imidazolium-based ILs (IILs) with varying anions namely dicyanamide (DCA), hydrogen sulfate (HS), dimethyl phosphate (DP), acetate (A), sulfate (S) and dihydrogen phosphate (DHP) have been chosen with the aim of understanding the role of anions in bringing about the destabilization effect on collagen based on the kosmotropicity and chaotropicity of ions. Imidazolium-based ILs destabilized the triple helical structure of collagen, thereby proving as strong denaturants for collagen and this was confirmed by various spectroscopic techniques viz., CD, FT-IR, viscosity and impedance measurements. The solution studies were in accordance to the changes in the dimensional stability of RTT collagen fibres at the fibrillar level. Imidazolium cations with varied anions have exhibited destabilizing effect on collagen in order of ions in Hofmeister series; IDP < IDHP < IA < IDCA < IS < IHS. Presumably, these notable effect and changes were facilitated by electrostatic interactions between the anions and amine functional groups of collagen.

Choline-Based Amino Acid ILs-Collagen Interaction: Enunciating Its Role in Stabilization/Destabilization Phenomena.

Given the potential of productive interaction between choline-based amino acid ionic liquids (CAAILs) and collagen, we investigated the role of four CAAILs, viz., choline serinate, threoninate, lysinate, and phenylalaninate, and the changes mediated by them in the structure of collagen at different hierarchical orderings, that is, at molecular and fibrillar levels. The rheological, dielectric behavior and the secondary structural changes signify the alteration in the triple helical structure of collagen at higher concentrations of CAAILs. A marginal swelling and slight decrease in the thermal stability of rat tail tendon collagen fibers were observed for choline serinate and threoninate, albeit distortions in banding patterns were noticed for choline lysinate and phenylalaninate, suggesting chaotropicity of the ions at the fibrillar level. This signifies the changes in the hydrogen-bonding environment of collagen with increasing concentrations of CAAILs, which could be due to competitive hydrogen bonding between the carbonyl group of amino acid ionic liquids and the hydroxyl groups of collagen.

Impact of Ionic Liquids on the Structure and Dynamics of Collagen.

The changes in the structure and dynamics of collagen treated with two different classes of ionic liquids, bis-choline sulfate (CS) and 1-butyl-3-methyl imidazolium dimethyl phosphate (IDP), have been studied at the molecular and fibrillar levels. At the molecular level, circular dichroic studies revealed an increase in molar ellipticity values for CS when compared with native collagen, indicating cross-linking, albeit pronounced conformational changes for IDP were witnessed indicating denaturation. The impedance was analyzed to correlate the conformational changes with the hydration dynamics of protein. Changes in the dielectric properties of collagen observed upon treatment with CS and IDP reported molecular reorientation in the surrounding water milieu, suggesting compactness or destabilization of the collagen. This was further confirmed by proton transverse NMR relaxation time measurements, which demonstrated that the water mobility changes in the presence of the ILs. At the fibrillar level, differential scanning calorimetry thermograms for rat tail tendon collagen fibers treated with CS show a 5 °C increase in denaturation temperature, suggesting imparted stability. On the contrary, a significant temperature decrease was noticed for IDP, indicating the destabilization of collagen fibers. The obtained results clearly indicate that the changes in the secondary structure of protein are due to the changes in the hydration dynamics of collagen upon interaction with ILs. Thus, this study on the interaction of collagen with ionic liquids unfolds the propensity of ILs to stabilize or destabilize collagen depending on the changes invoked at the molecular level in terms of structure and dynamics of protein, which also got manifested at the fibrillar level.

Role of Preferential Ions of Ammonium Ionic Liquid in Destabilization of Collagen.

Ions play a key role in the destabilization of collagen. This study explores the effect of diethyl methyl ammonium methane sulfonate (AMS), an ionic liquid (IL), on different hierarchical orderings of collagen, namely, at the molecular and fibrillar levels. The rheological behavior and secondary structural changes reveal changes in the hydrogen-bonding environment of collagen, leading to alterations in the triple helical structure of collagen. An increase in the concentration of AMS resulted in swelling of rat-tail tendon fibers, and also, decreased thermal stability signifies that ions are obliged to destabilize collagen at the fibrillar level. Molecular modeling studies confirm that anions are judiciously held responsible for structural deformities in collagen, whereas cations have a tenuous effect. Thus, the preferential role of ions present in an ammonium IL has been elucidated in this study.