International Journal of Biological Macromolecules 36 (2005) 39–46 Microscopic structure of gelatin coacervates Biswaranjan Mohanty, H.B. Bohidar ∗ School of Physical Sciences, Jawaharlal Nehru University, New Delhi 110067, India Received 16 August 2004; received in revised form 16 March 2005; accepted 21 March 2005 Available online 17 May 2005 Abstract Microscopic structure of simple coacervates of gelatin having concentration ∼130 g/l were studied at 25 ◦ C by atomic force microscopy (AFM), rheology, small angle neutron scattering (SANS), UV absorption and circular dichroism (CD) techniques. The behavior of viscoelastic exponents Δ ′ and Δ ′′ of storage and loss modulii (G ′ (ω) ∼ ω Δ ′ , G ′′ (ω) ∼ ω Δ ′′ ) revealed that, Δ ′ = 0.25 ± 0.01 and Δ ′′ = 0.78 ± 0.1 for coac- ervates. The mass fractal dimension ‘d f ’ for coacervate was found to be 2.27, which attributed a compact heterogeneous network structure to the coacervates. This is supported by AFM pictures. The CD and UV absorption data indicated presence of helical structures inside the coacervates phase. SANS results showed the existence of a single length scale associated with this system identified as gelatin persistence length,  = 27 ± 2 ˚ A. These studies indicate that the coacervate phase is a low dimensional dense heterogeneous material comprised of strongly interconnected triple helices which imparts a large storage modulus to this material. © 2005 Elsevier B.V. All rights reserved. Keywords: Coacervates; Rheology; Neutron scattering; Circular dichroism; Gelatin 1. Introduction A macromolecule can remain in equilibrium with its sol- vent at room temperature in three physically distinct states namely; solution [1–3], gel [4–9] and coacervates [10–12]. Gelatin forms physical gels in hydrogen bond friendly sol- vents above a concentration larger than the chain overlap concentration c ∗ (≈2%, w/v) that relates to the intrinsic vis- cosity [η] as c ∗ ∼ 1/[η]. The gelatin sol undergoes a first- order thermo-reversible gelation transition for temperatures T < T g , where T g (≈30 ◦ C) is the gelation temperature. Dur- ing this process the gelatin molecules undergo an associa- tion mediated conformational transition from random coil to triple helix [5–8]. The sol has polydisperse random coils of gelatin molecules and aggregates [1–3] whereas in the gel state there is a propensity of triple helices stabilized through intermolecular hydrogen bonding. During this pro- cess, a three-dimensional interconnected network connecting large fraction of the gelatin chains is formed [4,13]. ∗ Corresponding author. Tel.: +91 11 267 175 03; fax: +91 11 267 175 37. E-mail address: bohi0700@mail.jnu.ac.in (H.B. Bohidar). On the contrary very little is known about the structure of gelatin coacervates. In simple coacervation process a ho- mogeneous solution of 1% (w/v) gelatin is driven towards a liquid–liquid phase separation, giving rise to a polymer dense phase remaining in equilibrium with its supernatant. Coacervates are formed through sequential self-charge neu- tralization of gelatin molecules that are mostly intermolecu- lar. This is described in excellent details elsewhere [10,12]. Gelatin is a weak polyampholyte (with random coil persis- tence length ≈2.5 nm [13]) with typically the positive and negatively charged segments occurring in its backbone in the ratio 1:1 which constitute about 22% of chain length together. Addition of a non-solvent like alcohol facilitates the process of inducing chain collapse and the positively charged seg- ments interact with the negatively charged segments through screened Coulomb interactions, the details of this and phase diagrams are discussed elsewhere [10]. Gelatin based coacer- vates have shown promise in drug and enzyme encapsulation. Alcohol induced coacervation increases the shelf-life of coac- ervates which owes its origin to the bactericidal properties of alcohol. In gelatin coacervation studies, the coherent picture that had emerged was [10]: (i) that a homogeneous solution con- 0141-8130/$ – see front matter © 2005 Elsevier B.V. All rights reserved. doi:10.1016/j.ijbiomac.2005.03.012