Investigating the use of conducting oligomers and redox molecules in CdS-MoFeP biohybrids.

In this work we report the effect of incorporating conducting oligophenylenes and a cobaltocene-based redox mediator on photodriven electron transfer between thioglycolic acid (TGA) capped CdS nanorods (NR) and the native nitrogenase MoFe protein (MoFeP) by following the reduction of H+ to H2. First, we demonstrate that the addition of benzidine-a conductive diphenylene- to TGA-CdS and MoFeP increased catalytic activity by up to 3-fold as compared to CdS-MoFeP alone. In addition, in comparing the use of oligophenylenes composed of one (p-phenylenediamine), two (benzidine) or three (4,4''-diamino-p-terphenyl)phenylene groups, the largest gain in H2 was observed with the addition of benzidine and the lowest with phenylenediamine. As a comparison to the conductive oligophenylenes, a cobaltocene-based redox mediator was also tested with the TGA-CdS NRs and MoFeP. However, adding either cobaltocene diacid or diamine caused negligible gains in H2 production and at higher concentrations, caused a significant decrease. Agarose gel electrophoresis revealed little to no detectable interaction between benzidine and TGA-CdS but strong binding between cobaltocene and TGA-CdS. These results suggest that the tight binding of the cobaltocene mediator to CdS may hinder electron transfer between CdS and MoFe and cause the mediator to undergo continuous reduction/oxidation events at the surface of CdS.

Investigating Protein-Nanocrystal Interactions for Photodriven Activity.

We illustrate how intermolecular interactions facilitate ATP-free electron transfer between either native or engineered MoFe protein (MoFeP) from nitrogenase and a CdS nanorod (NR) by following the reduction of H+ to H2. First, by varying the charge on the surface of the NR, we show the role of electrostatic interactions on MoFeP binding to the particle surface and subsequent H+ reduction. Next, the role of strong, semicovalent thiol-CdS interactions was tested using free cysteines on the MoFeP. By blocking free cysteines, we show that the presence of free thiols on the protein has little to no influence on CdS binding and resultant photocatalytic activity. We next studied methods to covalently bind the protein to CdS by modifying the free cysteines with dibenzocyclooctyne (DBCO) and reacting the CdS NRs capped with a mixture of negatively charged thioglycolic acid and thiol-PEG3-azide ligands. As compared to that of the unmodified proteins, a 32.2 ± 1.5% and 61.7 ± 2.1% increase in H2 production was observed from MoFeP and C-MoFeP, respectively. At last, to test the effect of both charge and covalent tethering, positively charged cysteamine/azide CdS NRs were reacted with DBCO-modified C-MoFeP, which showed little improvement over native C-MoFeP alone under irradiation. These results show the importance of both electrostatic associations between the NR and protein and covalently tethering the protein to the semiconductor surface for enhanced electron transfer and photodriven activity.

Click Nucleic Acid Mediated Loading of Prodrug Activating Enzymes in PEG-PLGA Nanoparticles for Combination Chemotherapy.

The simultaneous delivery of multiple therapeutics to a single site has shown promise for cancer targeting and treatment. However, because of the inherent differences in charge and size between drugs and biomolecules, new approaches are required for colocalization of unlike components in one delivery vehicle. In this work, we demonstrate that triblock copolymers containing click nucleic acids (CNAs) can be used to simultaneously load a prodrug enzyme (cytosine deaminase, CodA) and a chemotherapy drug (doxorubicin, DOX) in a single polymer nanoparticle. CNAs are synthetic analogs of DNA comprised of a thiolene backbone and nucleotide bases that can hybridize to complementary strands of DNA. In this study, CodA was appended with complementary DNA sequences and fluorescent dyes to allow its encapsulation in PEG-CNA-PLGA nanoparticles. The DNA-modified CodA was found to retain its enzyme activity for converting prodrug 5-fluorocytosine (5-FC) to active 5-fluorouracil (5-FU) using a modified fluorescent assay. The DNA-conjugated CodA was then loaded into the PEG-CNA-PLGA nanoparticles and tested for cell cytotoxicity in the presence of the 5-FC prodrug. To study the effect of coloading DOX and CodA within a single nanoparticle, cell toxicity assays were run to compare dually loaded nanoparticles with nanoparticles loaded only with either DOX or CodA. We show that the highest level of cell death occurred when both DOX and CodA were simultaneously entrapped and delivered to cells in the presence of 5-FC.

Solar Photocatalytic Phenol Polymerization and Hydrogen Generation for Flocculation of Wastewater Impurities.

Achieving global sustainability will require balancing encroaching climate changes while maintaining existing quality of life. Using sunlight to purify wastewater while simultaneously generating usable fuels is an opportunity to approach both targets in a cost-efficient manner. In addition, converting biomass products to usable polymers is a sustainable approach for potentially replacing polystyrene or other petroleum derived polymers. Phenols from medical, manufacturing, and agricultural waste are commonly found in many water sources, and they are known to foul common reverse osmosis membranes. Here, we show oxidative polymerization of guaiacol, an aromatic compound derived from biomass, with concurrent hydrogen gas generation by using platinum-seeded cadmium sulfide nanorods (Pt@CdS) as photocatalysts. Rather than forming short oligomers as typically made by enzymes such as laccase and peroxidase, the resulting polymers show higher molecular weights that can more easily flocculate out of water. By comparing guaiacol conversion to molecular weight and dispersity, the guaiacol was found to polymerize via a chain-growth process. We also show that Pt@CdS can polymerize other phenols as well by testing the monomers phenol, 2,6-dihydroxybenzoic acid, gallic acid, and vanillin. Lastly, because the aqueous solubility of these aromatic polymers decreases dramatically with molecular weight, polymerization reactions were also tested in biphasic solutions to determine if chain growth could propagate in the oil phase. We show that the Pt@CdS nanoparticles can form stable Pickering emulsions in various biphasic combinations, and that both H2 formation and polymer molecular weight correlated with the partition coefficient of guaiacol into the oil phase as well as the solubility of the growing polymer chains. These combined studies demonstrate the possibility of using nanoscale photocatalysts to oxidatively polymerize phenolic substrates via a chain-growth mechanism, thereby providing a path for pretreating water by flocculating out contaminants with concurrent generation of hydrogen.

Multicatalytic, Light-Driven Upgrading of Butanol to 2-Ethylhexenal and Hydrogen under Mild Aqueous Conditions.

Microbes produce low-molecular-weight alcohols from sugar, but these metabolites are difficult to separate from water and possess relatively low heating values. A combination of photo-, organo-, and enzyme catalysis is shown here to convert C4 butanol (BuOH) to C8 2-ethylhexenal (2-EH) using only solar energy to drive the process. First, alcohol dehydrogenase (ADH) catalyzed the oxidation of BuOH to butyraldehyde (BA), using NAD+ as a cofactor. To prevent back reaction, NAD+ was regenerated using a platinum-seeded cadmium sulfide (Pt@CdS) photocatalyst. An amine-based organocatalyst then upgraded BA to 2-EH under mild aqueous conditions rather than harsh basic conditions in order to preserve enzyme and photocatalyst stability. The process also simultaneously increased total BuOH conversion. Thus, three disparate types of catalysts synergistically generated C8 products from C4 alcohols under green chemistry conditions of neutral pH, low temperature, and pressure.

Light-Driven Catalytic Upgrading of Butanol in a Biohybrid Photoelectrochemical System.

This paper reports the design and preparation of a biohybrid photoelectrochemical cell (PEC) that can drive the tandem enzymatic oxidation and aldol condensation of n-butanol (BuOH) to C8 2-ethylhexenal (2-EH). In this work, BuOH was first oxidized to n-butyraldehyde (BA) by the alcohol oxidase enzyme (AOx), concurrently generating hydrogen peroxide (H2O2). To preserve enzyme activity and increase kinetics nearly 2-fold, the H2O2 was removed by oxidation at a bismuth vanadate (BiVO4) photoanode. Organocatalyzed aldol condensation of C4 BA to C8 2-EH improved the overall BuOH conversion to 6.2 ± 0.1% in a biased PEC after 16 h. A purely light-driven, unbiased PEC showed 3.1 ± 0.1% BuOH conversion, or ~50% of that obtained from the biased system. Replacing AOx with the enzyme alcohol dehydrogenase (ADH), which requires the diffusional nicotinamide adenine dinucleotide cofactor (NAD+/NADH), resulted in only 0.2% BuOH conversion due to NAD+ dimerization at the photoanode. Lastly, the application of more positive biases with the biohybrid AOx PEC led to measurable production of H2 at the cathode, but at the cost of lower BA and 2-EH yields due to both product overoxidation and decreased enzyme activity.

Conversion of Ethanol to 2-Ethylhexenal at Ambient Conditions Using Tandem, Biphasic Catalysis.

Ethanol is a ubiquitous fermentation product well-tolerated by microbes, but purification from growth media requires multiple distillations or other energy intensive processes. Converting such metabolites to larger, hydrophobic products would both yield higher energy products and facilitate separation. Here, we demonstrate the conversion of C2 ethanol to C8 2-ethylhexenal via a sequential oxidation-aldol-hydrogenation-aldol process with solar energy as the only required input. Photocatalysis was utilized to drive enzymatic oxidation of ethanol, while biphasic media in conjunction with aldol coupling and Pd assisted hydrogenation kept the oxidation and reduction processes physically and chemically separated. Using this process, 2-ethylhexenal was produced from ethanol in both buffer and diluted yeast media.