Subhalakshmi Nagarajan

Biocatalytically Oligomerized Epicatechin with Potent and Specific Anti-proliferative Activity for Human Breast Cancer Cells

Catechins, naturally occurring flavonoids derived from wine and green tea, are known to exhibit multiple health benefits. Epigallocatechin gallate (EGCG) is one of the most widely investigated catechins, but its efficacy in cancer therapy is still inconsistent and limited. The poor stability of EGCG has contributed to the disparity in the reported anti-cancer activity and other beneficial properties. Here we report an innovative enzymatic strategy for the oligomerization of catechins (specifically epicatechin) that yields stable, water-soluble oligomerized epicatechins with enhanced and highly specific anti-proliferative activity for human breast cancer cells. This one-pot oxidative oligomerization is carried out in ambient conditions using Horseradish Peroxidase (HRP) as a catalyst yielding water-soluble oligo(epicatechins). The oligomerized epicatechins obtained exhibit excellent growth inhibitory effects against human breast cancer cells with greater specificity towards growth-inhibiting cancer cells as opposed to normal cells, achieving a high therapeutic differential. Our studies indicate that water-soluble oligomeric epicatechins surpass EGCG in stability, selectivity and efficacy at lower doses.

Halogen-free ultra-high flame retardant polymers through enzyme catalysis

Over the past few years, the use of certain types of halogenated flame retardant additives such as decabromodiphenylether has come under intense scrutiny due to their toxicity, environmental persistence and bio-accumulation. There is an immediate need for the development of non-toxic alternative flame retardant materials and fire resistant polymers with comparable or better efficacies, obtained using benign synthetic approaches. Enzymatic polymerization is being used increasingly as an environmentally friendly alternative method for the synthesis of functional materials including polymers and additives. Here, we report a biocatalytic synthesis of a new class of thermally stable, ultra-fire resistant polyphenols based on deoxybenzoins. In calorimetric studies, these polyphenols exhibit very low heat release capacities (comparable to Nomex™) and form a large amount of carbonaceous char rendering them suitable for flame retardant applications.

Enzymatic Synthesis and Characterization of PolyQuercetin

Quercetin is a flavonol found in fruits, onions and wine. Recently, quercetin has been extensively investigated for its antioxidant behavior in food such as poultry and fish. While quercetin has been shown to be a good stabilizer for several oils, the thermal stability and solubility at neutral and slightly alkaline conditions continue to be an area of huge concern. However, increasing the stability and the antioxidant potency of this flavonol would be beneficial to the food industry. We utilized horseradish peroxidase, a phytochemical enzyme, to polymerize quercetin in biocompatible water/ethanol mixtures. This unique, one pot procedure has provided a method to synthesize polyquercetin under mild conditions. The final polymer is soluble in a water/ethanol mixture. Preliminary data were presented in the Material Research Society fall meeting in Boston on December 3, 2008. However, in this article, enzymatically synthesized polyquercetin has also been characterized by a variety of techniques. Using UV-Vis FTIR, GPC, NMR and TGA we have deduced certain aspects of the structure. Structural elucidation was further refined by results from molecular modeling. Density functional theory calculations predict that the UV-Vis spectra of polyquercetin can show high conjugation relative to those of quercetin monomer. This was confirmed by the experimental results. This novel thermally stable polymer, synthesized using Green Chemistry principles, can be used as a potent antioxidant in the highly regulated food industry.

Biocatalytic Synthesis of Water-Soluble Oligo(catechins)

Catechins are polyphenolic compounds found in green tea. These flavonoids possess anti‐oxidant, anti‐carcinogenic, and anti‐inflammatory properties. The primary goal of this work is to synthesize water‐soluble oligomers of catechins. Although, oxidative polymerization of catechins catalyzed by enzymes such as Horseradish Peroxidase (HRP) have been reported, the polymeric products have limited solubility. This restricts their compatibility with biological systems and their effectiveness as antioxidants or anti‐carcinogenic agents. Here we report a unique enzymatic approach for the synthesis of water‐soluble oligo(catechins). Polyelectrolytes such as Sulfonated Polystyrene (SPS) and surfactants such as sodium dodecylbenzenesulphonate (SDS) have been used as templates for the HRP catalyzed polymerization of (+)‐, (−)‐catechins and (−)‐epicatechin. Oligocatechins synthesized exhibit enhanced anti‐tumorigenic activity to human colon cancer cells as compared with the monomeric catechins.

A stable biomimetic redox catalyst obtained by the enzyme catalyzed amidation of iron porphyrin

Hematin, a hydroxyferriprotoporphyrin, is the more stable oxidized form of the free heme. The use of hematin as a catalyst for oxidative polymerization reactions has been restricted due to its limited aqueous solubility at low pH conditions. While there have been reports on the functionalization of hematin with poly (ethylene glycol), the esters formed are not very stable at low pH conditions. We report here the design and synthesis of hematin tethered with methoxypolyethylene glycol amine chains as a novel stable biomimetic catalyst. This one step amidation was performed under solventless conditions and catalyzed by a hydrolase (Novozyme-435). The amidation greatly improved the stability of hematin at low pH. Further, this catalyst was soluble in water and was able to catalyze the polymerization of aniline based monomers. The amide functionalized hematin serves as a robust cost-effective alternative to HRP, active even at lower pH conditions.

Biocatalytic Modification of Naturally Occurring Iron Porphyrin

Hematin, a hydroxyferriprotoporphyrin, is the stable, oxidized form of heme. Heme has been reported to be the active catalytic center of naturally occurring peroxidases such as horseradish peroxidase (HRP). While there have been reports on the use of hematin as a catalyst for oxidative polymerization reactions, these reactions could be carried out only at high pH conditions due to limited aqueous solubility of hematin at lower pH conditions. We report here the biocatalytic modification of hematin using a lipase, Novozyme-435. Hematin has been modified by tethering monomethoxy polyethylene glycol (mPEG) chains which provide aqueous solubility over a fairly wide range of pH conditions. This pegylated Hematin (PEG-Hematin) is synthesized via a one-step solventless reaction and the products formed can be isolated with minimal purification. The PEG-Hematin synthesized serves as a robust alternative to HRP for the polymerization of aniline and phenol.

A Simple Technique for Submicron Scale Patterning of Silver Using Visible Light Interference

A novel, one step and simple methodology for the fabrication of submicron scale silver patterns is demonstrated. The photosensitivity of an organic silver salt has been utilized for this purpose of fabrication. The silver-organometallic compound is converted to metallic silver selectively in the illuminated regions. Surface morphology was studied by scanning electron microscopy (SEM). Energy dispersion spectroscopy (EDS) shows the presence of silver in the developed film. X-ray photoelectron spectroscopy (XPS) confirms the formation of metallic silver. Feature sizes of the order of 200 nm have been achieved using this technique.

Detection of Explosives using nanofibrous membranes

We report an inexpensive and simple approach for the fabrication of high surface area sensing elements using electrospinning. The nanofibrous membranes formed could be used as chemiresistors as well as fluorescence quenching sensors depending on the receptor used. For chemiresistor type sensors, conducting polymer (CP) nanotubes have been fabricated by combining electrospinning and vapor deposition polymerization. UV-Visible spectroscopy and X-ray photoelectron spectroscopy (XPS) confirm the formation of CP on the fiber. For an optical response, fluorescent dyes have been incorporated on the surface of the nanofibers. The quenching of fluorescence typically follows a Stern-Volmer type bimolecular quenching relationship. The morphological characterization was carried out using scanning electron microscopy (SEM). The response of these sensors to an explosive stimulant will be discussed.

Biocatalytic Synthesis of Two-Photon Active Resveratrol Oligomer

Resveratrol (3,5,4′ trihydroxy trans-stilbene) is a plant based phenolic compound. Enzymatic oligomerization of trans-resveratrol using horseradish peroxidase followed by characterization of the oligomer is presented. The oligomerization reaction was monitored using UV-Visible absorption and fluorescence spectroscopies. The oligomer exhibits strong two-photon-induced fluorescence. Computational modeling using spin-density calculations was performed to investigate the most probable reaction sites and the nature of products formed in the oligomerization process.

Biocatalysis for Material Science and Drug Discoveries

A novel biomimetic route for the synthesis of conducting homopolymers and copolymers from aniline, phenol, pyrrole and 3,4-ethylenedioxy-thiophene in the presence of a polyelectrolyte, such as polystyrene sulfonate (SPS) is presented. A poly(ethylene glycol) modified hematin (PEG-Hematin) and the enzyme horseradish peroxidase (HRP) were used to catalyze the copolymerization of different monomers. UV-vis, FTIR, XPS, TGA and electrical conductivity studies for all complexes indicated the presence of a stable and electrically conductive form of these polymers. Furthermore, the presence of a polyelectrolyte, such as SPS, in this complex provides a unique combination of properties such as processability and water-solubility. Additionally catechins, the active compounds found in green tea, were polymerized and found to exhibit very interesting anti-carcinogenic properties. Here we report a unique enzymatic approach for the synthesis of water-soluble poly(catechins) with enhanced stability and potent anti-proliferative effects on human cancer cells in vitro. Various stereoisomers of catechin [(+), (-), (±)] and (-)-epicatechin have been biocatalytically polymerized using HRP in ethanol/buffer mixtures. This one-pot biocatalytic polymerization is carried out in ambient conditions yielding water-soluble poly(catechins). These synthesized poly(catechins) were tested for their growth inhibitory properties using a variety of normal and cancerous human epithelial cell lines. The poly(catechins) exhibit statistically significant greater growth inhibitory effects when compared to the monomers and exhibited specificity, inhibiting the growth of breast, colorectal and esophageal cancer cells while having little effect on normal epithelial cell growththus achieving a high therapeutic ratio. The synthesis, characterization and the growth inhibitory effects of these novel water-soluble poly(catechins) will also be presented.