Determination of efficacy of a novel alginate dressing in a lethal arterial injury model in swine.

INTRODUCTION: Alginate is a biocompatible polysaccharide that is commonly used in the pharmaceutical, biomedical, cosmetic, and food industries. Though solid dressings composed of alginate can absorb water and promote wound healing, they are not effective hemostatic materials, particularly against massive hemorrhage. The purpose of this study is to attempt to increase the hemostatic capabilities of alginate by means of hydrophobic modification. Previous studies have illustrated that modifying a different polysaccharide, chitosan, in this way enhances its hemostatic efficacy as well as its adhesion to tissue. Here, it was hypothesized that modifying alginate with hydrophobic groups would demonstrate analogous effects. METHODS: Fifteen Yorkshire swine were randomized to receive hydrophobically-modified (hm) alginate lyophilized sponges (n=5), unmodified alginate lyophilized sponges (n=5), or standard Kerlix™ gauze dressing (n=5) for hemostatic control. Following a splenectomy, arterial puncture (6mm punch) of the femoral artery was made. Wounds were allowed to freely bleed for 30s, at which time dressings were applied and compressed for 3min in a randomized fashion. Fluid resuscitation was given to preserve the baseline mean arterial pressure. Wounds were monitored for 180min after arterial puncture, and surviving animals were euthanized. RESULTS: Blood loss for the hm-alginate group was significantly less than the two control groups of (1) alginate and (2) Kerlix™ gauze (p=<0.0001). Furthermore, 80% of hm-alginate sponges were able to sustain hemostasis for the full 180min, whereas 0% of dressings from the control groups were able to achieve initial hemostasis. CONCLUSIONS: Hm-alginate demonstrates a greatly superior efficacy, relative to unmodified alginate and Kerlix™ gauze dressings, in achieving hemostasis from a lethal femoral artery puncture in swine. This is a similar result as has been previously described when performing hydrophobic modification to chitosan. The current study further suggests that hydrophobic modification of a hydrophilic biopolymer backbone can significantly increase the hemostatic capabilities relative to the native biopolymer.

Light-Directed Self-Assembly of Robust Alginate Gels at Precise Locations in Microfluidic Channels.

Recently there has been much interest in using light to activate self-assembly of molecules in a fluid, leading to gelation. The advantage of light over other stimuli lies in its spatial selectivity, i.e., its ability to be directed at a precise location, which could be particularly useful in microfluidic applications. However, existing light-responsive fluids are not suitable for these purposes since they do not convert into sufficiently strong gels that can withstand shear. Here, we address this deficiency by developing a new light-responsive system based on the well-known polysaccharide, alginate. The fluid is composed entirely of commercially available components: alginate, a photoacid generator (PAG), and a chelated complex of divalent strontium (Sr(2+)) cations. Upon exposure to ultraviolet (UV) light, the PAG dissociates to release H(+) ions, which in turn induce the release of free Sr(2+) from the chelate. The Sr(2+) ions self-assemble with the alginate chains to give a stiff gel with an elastic modulus ∼2000 Pa and a yield stress ∼400 Pa (this gel is strong enough to be picked up and held by one's fingers). The above fluid is sent through a network of microchannels and a short segment of a specific channel is exposed to UV light. At that point, the fluid is locally transformed into a strong gel in a few minutes, and the resulting gel blocks the flow through that channel while other channels remain open. When the UV light is removed, the gel is gradually diluted by the flow and the channel reopens. We have thus demonstrated a remote-controlled fluidic valve that can be closed by shining light and reopened when the light is removed. In addition, we also show that light-induced gelation of our alginate fluid can be used to deposit biocompatible payloads at specific addresses within a microchannel.

Reversible gelation of cells using self-assembling hydrophobically-modified biopolymers: towards self-assembly of tissue.

Polymer hydrogels have long been used to hold and culture biological cells within their three-dimensional (3-D) matrices. Typically, in such cases, the cells are passively entrapped in a mesh of polymer chains. Here, we demonstrate an alternate approach where cells serve as active structural elements (crosslinks) within a polymer gel network. The polymers used in this context are hydrophobically modified (hm) derivatives of common biopolymers such as chitosan and alginate. We show that hm-polymers rapidly transform a liquid suspension of cells into an elastic gel. In contrast, the native biopolymers (without hydrophobes) do not cause such gelation. Gelation occurs because the hydrophobes on the polymer get embedded within the hydrophobic interiors of cell bilayer membranes. The polymer chains thus connect the cells into a 3-D sample-spanning network, with the cells serving as the junctions in the network. We demonstrate that a variety of cell types, including blood cells, endothelial cells, and breast cancer cells can be gelled by this approach. Cells gelled by hm-alginate are shown to remain viable within the network. Also, since the crosslinking mechanism is based on hydrophobic interactions, we show that the addition of supramolecules with hydrophobic binding pockets can reverse the gelation and release the cells. Cell-gels can be employed as injectable biomaterials since the connections in the network are susceptible to shear, but recover rapidly once shear is stopped. The overall approach provides a simple route towards the directed assembly of cell clusters and potentially to living tissue.

Gelation of vesicles and nanoparticles using water-soluble hydrophobically modified chitosan.

Hydrophobically modified chitosan (hmC) is a self-assembling polymer that has attracted recent attention for many applications, including as a hemostatic agent. One limitation with chitosan and its derivatives like hmC is that these polymers are soluble in water only under acidic conditions (because the pKa of chitosan is about 6.5), which could be undesirable for biomedical applications. To circumvent this limitation, we have synthesized a derivative of a C12-tailed hmC that is soluble in water at neutral pH. This water-soluble hmC (ws-hmC) is obtained by grafting O-carboxymethyl groups onto some of the primary hydroxyls on hmC. The solubility of ws-hmC at neutral pH is shown to be the result of a net anionic character for the polymer due to ionization of the carboxymethyl groups (in comparison, hmC is cationic). We also demonstrate that ws-hmC retains the self-assembling properties of hmC. Specifically, ws-hmC is able to induce gelation at neutral pH in dispersions of anionic surfactant vesicles as well as polymethylmethacrylate latex nanoparticles. Gelation is attributed to hydrophobic interactions between the hydrophobes on ws-hmC with vesicle bilayers and nanoparticle surfaces. In each case, gelation can be reversed by the addition of α-cyclodextrin, a supramolecule with a hydrophobic cavity that sequesters the hydrophobes on the polymer.

Glucose oxidase-mediated gelation: a simple test to detect glucose in food products.

This paper reports a simple, rapid, and sugar-selective method to induce gelation from glucose-containing samples. This method employs glucose oxidase (GOx) to selectively "recognize" and oxidize glucose to generate gluconic acid, which acts to solubilize calcium carbonate and release calcium ions. The release of calcium ions triggers gelation of the calcium-responsive polysaccharide alginate to form a calcium-alginate hydrogel. Rheological measurements confirm that gel formation is triggered by glucose but not fructose or sucrose (consistent with GOx's selectivity). Vial inversion tests demonstrate that gel formation can be readily observed without the need for instrumentation. Proof-of-concept studies demonstrate that this gel-forming method can detect glucose in food/beverage products sweetened with glucose or high-fructose corn syrups. These results indicate that the enzyme-induced gelation of alginate may provide a simple means to test for sweeteners using components that are safe for use on-site or in the home.

Light-activated ionic gelation of common biopolymers.

Biopolymers such as alginate and pectin are well known for their ability to undergo gelation upon addition of multivalent cations such as calcium (Ca(2+)). Here, we report a simple way to activate such ionic gelation by UV irradiation. Our approach involves combining an insoluble salt of the cation (e.g., calcium carbonate, CaCO(3)) with an aqueous solution of the polymer (e.g., alginate) along with a third component, a photoacid generator (PAG). Upon UV irradiation, the PAG dissociates to release H(+) ions, which react with the CaCO(3) to generate free Ca(2+). In turn, the Ca(2+) ions cross-link the alginate chains into a physical network, thereby resulting in a hydrogel. Dynamic rheological experiments confirm the elastic character of the alginate gel, and the gel modulus is shown to be tunable via the irradiation time as well as the PAG and alginate concentrations. The above approach is easily extended to other biopolymers such as pectin. Using this approach, a photoresponse can be imparted to conventional biopolymers without the need for any chemical modification of the molecules. Photoresponsive alginate gels may be useful in creating biomaterials or tissue mimics. As a step toward potential applications, we demonstrate the ability to photopattern a thin film of alginate gel onto a glass substrate under mild conditions.