A versatile method for preparation of hydrated microbial-latex biocatalytic coatings for gas absorption and gas evolution.

We describe a latex wet coalescence method for gas-phase immobilization of microorganisms on paper which does not require drying for adhesion. This method reduces drying stresses to the microbes. It is applicable for microorganisms that do not tolerate desiccation stress during latex drying even in the presence of carbohydrates. Small surface area, 10-65 µm thick coatings were generated on chromatography paper strips and placed in the head-space of vertical sealed tubes containing liquid to hydrate the paper. These gas-phase microbial coatings hydrated by liquid in the paper pore space demonstrated absorption or evolution of H2, CO, CO2 or O2. The microbial products produced, ethanol and acetate, diffuse into the hydrated paper pores and accumulate in the liquid at the bottom of the tube. The paper provides hydration to the back side of the coating and also separates the biocatalyst from the products. Coating reactivity was demonstrated for Chlamydomonas reinhardtii CC124, which consumed CO2 and produced 10.2 ± 0.2 mmol O2 m⁻² h⁻¹, Rhodopseudomonas palustris CGA009, which consumed acetate and produced 0.47 ± 0.04 mmol H2 m⁻² h⁻¹, Clostridium ljungdahlii OTA1, which consumed 6 mmol CO m⁻² h⁻¹, and Synechococcus sp. PCC7002, which consumed CO2 and produced 5.00 ± 0.25 mmol O2 m⁻² h⁻¹. Coating thickness and microstructure were related to microbe size as determined by digital micrometry, profilometry, and confocal microscopy. The immobilization of different microorganisms in thin adhesive films in the gas phase demonstrates the utility of this method for evaluating genetically optimized microorganisms for gas absorption and gas evolution.

Uniform lab-scale biocatalytic nanoporous latex coatings for reactive microorganisms.

This chapter describes a method for generating uniform lab-scale biocatalytic nanoporous latex coatings. Nearly everything we come into contact with on a daily basis has been coated with some polymer material. High-speed waterborne polymer coating and ink-jet printing techniques are mature technologies. Methods for immobilizing microorganisms in lab-scale waterborne latex biocatalytic coatings draw on existing coating technologies for generating precision industrial paint and paper coatings and would therefore be amenable to scale up in future applications. An inherent problem for many lab-scale techniques is coating uniformity. The method described here has been developed to dramatically increase the uniformity of multiple individual small surface area coatings derived from a single coating template by minimizing edge effects due to emulsion drying adjacent to the edge of the mask.

Progress toward a biomimetic leaf: 4,000 h of hydrogen production by coating-stabilized nongrowing photosynthetic Rhodopseudomonas palustris.

Intact cells are the most stable form of nature's photosynthetic machinery. Coating-immobilized microbes have the potential to revolutionize the design of photoabsorbers for conversion of sunlight into fuels. Multi-layer adhesive polymer coatings could spatially combine photoreactive bacteria and algae (complementary biological irradiance spectra) creating high surface area, thin, flexible structures optimized for light trapping, and production of hydrogen (H(2)) from water, lignin, pollutants, or waste organics. We report a model coating system which produced 2.08 +/- 0.01 mmol H(2) m(-2) h(-1) for 4,000 h with nongrowing Rhodopseudomonas palustris, a purple nonsulfur photosynthetic bacterium. This adhesive, flexible, nanoporous Rps. palustris latex coating produced 8.24 +/- 0.03 mol H(2) m(-2) in an argon atmosphere when supplied with acetate and light. A simple low-pressure hydrogen production and trapping system was tested using a 100 cm(2) coating. Rps. palustris CGA009 was combined in a bilayer coating with a carotenoid-less mutant of Rps. palustris (CrtI(-)) deficient in peripheral light harvesting (LH2) function. Cryogenic field emission gun scanning electron microscopy (cryo-FEG-SEM) and high-pressure freezing were used to visualize the microstructure of hydrated coatings. A light interaction and reactivity model was evaluated to predict optimal coating thickness for light absorption using the Kubelka-Munk theory (KMT) of reflectance and absorptance. A two-flux model predicted light saturation thickness with good agreement to observed H(2) evolution rate. A combined materials and modeling approach could be used for guiding cellular engineering of light trapping and reactivity to enhance overall photosynthetic efficiency per meter square of sunlight incident on photocatalysts.

Hydrogen production by photoreactive nanoporous latex coatings of nongrowing Rhodopseudomonas palustris CGA009.

Nonuniform light distribution is a fundamental limitation to biological hydrogen production by phototrophic bacteria. Numerous light distribution designs and culture conditions have been developed to reduce self-shading and nonuniform reactivity within bioreactors. In this study, highly concentrated (2.0 x 108 CFU/muL formulation) nongrowing Rhodopseudomonas palustris CGA009 were immobilized in thin, nanoporous, latex coatings. The coatings were used to study hydrogen production in an argon atmosphere as a function of coating composition, thickness, and light intensity. These coatings can be generated aerobically or anaerobically and are more reactive than an equivalent number of suspended or settled cells. Rhodopseudomonas palustris latex coatings remained active after hydrated storage for greater than 3 months in the dark and over 1 year when stored at -80 degrees C. The initial hydrogen production rate of the microphotobioreactors containing 6.25 cm2, 58.4 mum thick Rps. palustris latex coatings illuminated by 34.1 PAR mumol photons m-2 s-1 was 6.3 mmol H2 m-2 h-1 and had a final yield of 0.55 mol H2 m-2 in 120 h. A dispersible latex blend has been developed for direct comparison of the specific activity of settled, suspended, and immobilized Rps. palustris.